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Biomedical subjects

J Hardin

Publications and source records attributed to J Hardin.

At least 55 records · Page 3Linked to original sources
PubMed1991

Biosynthesis of human fibroblast growth factor-5.

We have analyzed the biosynthesis of human fibroblast growth factor-5 (FGF-5) at the translational and posttranslational levels. FGF-5 RNA synthesized in vitro can be translated in rabbit reticulocyte lysates to yield a 29,500-Da protein, which is consistent with the molecular weight predicted from the coding sequence. The efficiency of FGF-5 translation is dramatically enhanced if an upstream open reading frame (ORF-1) in the RNA is deleted or if both AUG codons in ORF-1 are destroyed by point mutations, while partial enhancement is achieved by individual mutation of either ORF-1 AUG codon. These data suggest that FGF-5 synthesis requires the scanning of ribosomes past the two ORF-1 AUG codons. The introduction of these ORF-1 mutations into a eukaryotic FGF-5 expression vector increases its capacity to transform mouse NIH 3T3 cells up to 50-fold upon transfection. FGF-5 is secreted from transfected 3T3 cells and from human tumor cells as glycoproteins containing heterogeneous amounts of sialic acid. Glycosidase treatments suggest that the growth factor bears both N-linked and O-linked sugars.

Amino Acid Sequence↗
PubMed1991

Epidermal growth factor upregulates intestinal electrolyte and nutrient transport.

The role of epidermal growth factor (EGF) in the acute regulation of intestinal transport of electrolytes and glucose was examined. In vivo transport studies were performed in New Zealand White rabbits (500-900 g) using a single-pass perfusion technique. In vitro net fluxes were determined under short-circuited conditions in Ussing chambers. Kinetic parameters of glucose transport in the presence and absence of EGF were measured in brush-border membrane vesicles. In vivo studies showed that the addition of 60 ng/ml EGF to the perfusate resulted in increased absorption of H2O, Na+, Cl-, and glucose from the jejunum. In Ussing chambers, the presence of EGF caused an increase in jejunal net fluxes of glucose-stimulated Na+ and 3-O-methylglucose due to an increase in mucosal-to-serosal movements. Verapamil abolished the EGF effect. In the absence of glucose, net fluxes of Na+ and Cl- were enhanced in the presence of EGF due to a decrease in the serosal-to-mucosal movement of both ions. Verapamil had no effect on this decrease. The incubation of EGF with brush-border membrane vesicles had no effect on either the maximal flux or the Michaelis constant of glucose transport. These results indicate that EGF is capable of regulating absorption of electrolytes and nutrients from the small intestine and suggest a role for this peptide in the control of intestinal transport.

Animals↗
PubMed1990

Target recognition by the archenteron during sea urchin gastrulation.

During sea urchin gastrulation filopodia are sent out by secondary mesenchyme cells (SMCs) at the tip of the archenteron in continual cycles of extension, attachment, and retraction. Eventually the archenteron ceases its elongation and its tip localizes to the animal pole region of the embryo (Gustafson and Kinnander, 1956, Exp. Cell Res. 11, 36-57; Dan and Okazaki, 1956, Biol. Bull. 110, 29-42). We have investigated the mechanisms and specificity of this localization by analyzing filopodial behavior and by experimental manipulation of the interaction of the archenteron with the animal pole region. When the tip of the archenteron nears the animal pole, some filopodia make contact with a well-defined locus within this region. Filopodia that make contact with the locus remain attached 20-50 times longer than attachments observed at any other site along the blastocoel wall. The SMCs bearing the long-lived filopodia eventually change their phenotype by flattening and spreading onto this region. Several lines of experimental evidence indicate that contact with the animal pole locus, or "target" region, is crucial for the change in phenotype of the SMCs: (1) the phenotypic change can be induced precociously by bringing the animal pole region within reach of the tip of the archenteron early in gastrulation. Precocious contact with other regions of the blastocoel wall does not induce a similar change. (2) The phenotypic change can be delayed by placing the animal pole out of reach late in gastrulation, resulting in artificial prolongation of exploratory behavior by filopodia. (3) Ectopic combinations of animal pole ectoderm and archenterons in fused multiple embryos and chimaeras result in attachment of archenterons to the nearest available target, and (4) freely migrating SMCs are observed to migrate randomly within the blastocoel, then stop at the animal pole and undergo the change in phenotype. Filopodia rapidly attach to the animal pole when the shape of early gastrulae is altered such that the animal pole is less than 35 microns from the tip of the archenteron, even though such attachments only occur in normal embryos at the 2/3-3/4 gastrula stage. Since it has previously been shown that the archenteron elongates autonomously to 2/3 of its final length (Hardin, 1988, Development 103, 317-324), it appears that autonomous extension of the archenteron is required to place filopodia close enough to the animal pole to allow them to interact with it.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1989

Higher order structure is present in the yeast nucleus: autoantibody probes demonstrate that the nucleolus lies opposite the spindle pole body.

A panel of sera from 892 autoimmune patients was screened by indirect immunofluorescence on mammalian cells. Seventy-three sera were identified that recognize the nucleolus. Three of these sera appear to stain the nucleolus in yeast, suggesting that they recognize highly conserved antigens. These three sera also immunoprecipitate mammalian U3 snRNA-containing particles, which reside in the nucleolus and have been implicated in rRNA processing. Double immunofluorescence experiments with anti-nucleolus and anti-tubulin antibodies revealed a novel form of non-random nuclear organization in yeast. The spindle pole body and the nucleolus-both of which are associated with the nuclear envelope-preferentially localize at opposite ends of the nucleus. Organization of these and other components into specific regions of the nucleus may be important for optimizing their proper function.

Autoantibodies↗
PubMed1989

Local shifts in position and polarized motility drive cell rearrangement during sea urchin gastrulation.

This study examines the mechanisms of epithelial cell rearrangement during archenteron elongation in the sea urchin embryo using scanning electron microscopy, differential interference contrast videomicroscopy, cell marking, and fluorescently labeled chimaeric clones. Archenteron elongation involves two major processes: local shifts in position of cells in the archenteron wall and polarized motility of the cells as they rearrange. Fluorescently labeled chimaeric clones introduced into the archenteron of Lytechinus pictus are initially 4-5 cells wide; by the end of gastrulation the clones elongate and narrow, so that they are one cell wide in the narrowest region of the archenteron. The extent of clonal mixing indicates that cells in the archenteron change their relative positions by only 1-2 cell diameters during cell rearrangement. Cells at the blastopore rearrange concomitantly with cells in the archenteron, resulting in a 35% decrease in blastopore diameter. Endoderm cells undergo polarized, stage-specific changes in shape and motility as they rearrange; (1) they flatten markedly along their apical-basal axis throughout archenteron elongation; (2) just prior to the onset of cell rearrangement, basal surfaces of all cells in the archenteron extend long, polarized lamellipodial protrusions along the axis of extension of the archenteron; (3) as cell rearrangement begins, basal surfaces round up and the cells become isodiametric; (4) by the 3/4 gastrula stage the cells become stretched along the animal-vegetal axis, apparently due to filopodial traction, and finally (5) they continue to rearrange, returning to a less elongated shape by the end of gastrulation. Direct observation of gastrulation in the cidaroid Eucidaris tribuloides indicates that in this species cell rearrangement is accomplished by progressive circumferential intercalation of cells without upwardly directed filopodia. This intercalation is accompanied by explosive, apparently stochastic, cortical blebbing activity at the boundaries between cells, suggesting that in addition to whatever cell rearrangement may be generated by filopodial tension, such activity is an important component of the active rearrangement process.

Animals↗
PubMed1989

Effect of Yersinia enterocolitica on intestinal mucin secretion.

Mucin and glycoprotein synthesis and secretion were evaluated in the upper, mid, and distal small intestine and in the proximal colon of rabbits infected with Yersinia enterocolitica (YE). Infected (INF) animals were examined on day 6 and compared with pair-fed controls and unmanipulated weight-matched rabbits. Tissue mucin content in vivo and mucin secretion in vitro, measured by a specific immunoassay, were significantly elevated in all four regions of the gut of INF rabbits compared with both control groups. In vitro secretion of stored glycoprotein, prelabeled with [3H]glucosamine, was not increased in the upper and mid small intestine of INF animals but was significantly elevated in the distal small intestine and proximal colon. In vitro incorporation of [14C]glucosamine was increased in all four regions of the gut of INF rabbits, but secretion of newly synthesized [14C]glycoprotein was only significantly elevated in the distal small intestine and proximal colon. A graded response was observed down the intestinal tract of INF rabbits, with the greatest increase in mucin content, synthesis and secretion occurring in the distal small intestine and proximal colon where the morphological impact of disease is also most severe.

Animals↗
PubMed1988

The behaviour and function of bottle cells during gastrulation of Xenopus laevis.

The behaviour of bottle cells in normal and microsurgically altered gastrulae and in cultured explants of Xenopus laevis was analysed, using time-lapse micrography, scanning electron microscopy (SEM) and cell tracing with fluorescein dextran amine (FDA). The results shed new light on the function of bottle cells. Bottle cells forming in vivo show a predominantly animal-vegetal apical contraction and a concurrent apical-basal elongation, whereas those forming in cultured explants show uniform apical contraction and remain rotund. Bottle cells forming in embryos with fewer subblastoporal cells contract more uniformly than those in normal embryos and release of normal bottle cells from supra- and subblastoporal cells results in immediate loss of the bottle shape. These results, and an analysis of the effects of bottle cell formation on the shapes and movements of surrounding tissues, show that unique shape of bottle cells and their probable function in development are not intrinsic properties but result from a modulation of the effect of a uniform and intrinsic apical contraction by the geometric and mechanical properties of the surrounding tissue. Mechanical simulations of bottle cell formation, using the finite element method, suggest how the site of bottle cell formation and the thickness and stiffness of adjacent tissues might change the effects of their formation. These results and FDA marking of prospective bottle cells and the adjacent deep mesodermal cells suggest that bottle cells function during their formation to initiate the involution of the prospective mesodermal mantle. Later they respread to deepen the archenteron and to form its peripheral wall.

Animals↗
PubMed1988

The role of secondary mesenchyme cells during sea urchin gastrulation studied by laser ablation.

It has long been thought that traction exerted by filopodia of secondary mesenchyme cells (SMCs) is a sufficient mechanism to account for elongation of the archenteron during sea urchin gastrulation. The filopodial traction hypothesis has been directly tested here by laser ablation of SMCs in gastrulae of the sea urchin, Lytechinus pictus. When SMCs are ablated at the onset of secondary invagination, the archenteron doubles in length at the normal rate of elongation, but advance of the tip of the archenteron stops at the 2/3 gastrula stage. In contrast, when all SMCs are ablated at or following the 2/3 gastrula stage, further elongation does not occur. However, if a few SMCs are allowed to remain in 2/3-3/4 gastrulae, elongation continues, although more slowly than in controls. The final length of archenterons in embryos ablated at the 1/3-1/2 gastrula stage is virtually identical to the final length of everted archenterons in LiCl-induced exogastrulae; since filopodial traction is not exerted in either case, an alternate, common mechanism of elongation probably operates in both cases. These results suggest that archenteron elongation involves two processes: (1) active, filopodia-independent elongation, which depends on active cell rearrangement and (2) filopodia-dependent elongation, which depends on mechanical tension exerted by the filopodia.

Animals↗
PubMed1987

The effect of perfluoropropane on the cornea in rabbits and cats.

The effects of perfluoropropane gas on the cornea were compared with those of sulfur hexafluoride gas following injection into the anterior chamber of rabbits and cats. Injection of 0.15 mL of gas produced a perfluoropropane bubble that lasted 22 days compared with a sulfur hexafluoride bubble that lasted seven days. The sulfur hexafluoride bubble produced corneal edema for as long as the gas was present. With perfluoropropane, corneal edema persisted even after the gas bubble disappeared. In another group of cats, multiple sequential injections of sulfur hexafluoride to maintain a gas bubble of duration comparable with the perfluoropropane bubble also produced edema as long as gas was present. Clinical corneal edema, endothelial fibrin deposition, endothelial opacities, and retrocorneal membrane were observed by slit-lamp and light, scanning, and transmission electron microscopic examinations of the corneas exposed to perfluoropropane but not of those exposed to sulfur hexafluoride. Prolonged corneal endothelial contact by perfluoropropane results in corneal edema due to endothelial dysfunction, which persists in cats probably because feline endothelium is less capable of regeneration than that of rabbit.

Animals↗
PubMed1987

The role of lysyl oxidase and collagen crosslinking during sea urchin development.

Lysyl oxidase, the only enzyme involved in collagen crosslinking, is shown to be present in embryos of the sea urchin Strongylocentrotus purpuratus. The enzyme specific activity increases over six-fold during development, showing the greatest rise during gastrulation and prism larva formation. The enzyme is inhibited by the specific inhibitor, beta-aminoproprionitrile (BAPN). Continuous BAPN treatment of S. purpuratus and Lytechinus pictus embryos from late cleavage stages onward increases the amount of noncrosslinked collagen present in prism larvae. When BAPN is added at the 128- or 256-cell stage it causes developmental arrest at the mesenchyme blastula stage. Embryos can be maintained in the arrested state for at least 96 h and will resume normal development and morphogenesis following BAPN removal. If BAPN is added after the mesenchyme blastula stage, it has little adverse effect on development; consequently nonspecific toxic effects of the drug are unlikely. The results suggest that lysyl oxidase and collagen crosslinking play a vital role in primary mesenchyme migration, gastrulation, and morphogenesis during sea urchin development and indicate that BAPN may be very useful in studying the extracellular matrix-cell interactions at the cellular and molecular level.

Amino Acid Oxidoreductases↗
PubMed1987

Cell behaviour during active cell rearrangement: evidence and speculations.

The cell behaviour and motility underlying cell rearrangement during gastrulation in amphibian and sea-urchin embryos are discussed. In particular, the cell behaviour of deep (non-epithelial) and epithelial cell populations that undergo cell rearrangement is compared and contrasted. Deep cell rearrangement in Xenopus laevis involves both convergence of cells towards the future dorsal midline and simultaneous axial extension of the mesodermal cell mass. Time-lapse cinemicrography and scanning electron microscopy suggest that asynchronous, repetitive motions of individual deep cells, involving local extensions and retractions of their margins, may provide the motive force for rearrangement. Such protrusive activity may be guided by local differences in cell-cell contacts in the marginal zone. Epithelial cell rearrangement in the sea-urchin embryo both elongates the archenteron and simultaneously closes the blastopore. Cell rearrangement is accompanied by stage-specific changes in protrusive activity and cell shape of the basal surfaces of cells in the wall of the gut rudiment, in contrast to the apical surfaces, which show little activity. These basal protrusions may be involved in the rearrangement process.

Animals↗
PubMed1986

The effects of aphidicolin on morphogenesis and differentiation in the sea urchin embryo.

Aphidicolin, an inhibitor of DNA polymerase alpha, blocks DNA synthesis and cell division in sea urchin embryos. The effects of this inhibition appear to be stage dependent. Blastulae treated with aphidicolin before the thickening of the vegetal plate undergo developmental arrest prior to gastrulation. The extent of inhibition of DNA synthesis varies from 60 to 93% in these embryos. However, when aphidicolin is added after the vegetal plate has thickened, development continues normally through pluteus formation, even though DNA synthesis is inhibited by greater than or equal to 90% and cell division has ceased. These observations indicate that, from the vegetal plate stage onward, morphogenesis and overt differentiation are independent of DNA synthesis and cell division.

Animals↗
PubMed1980

Humoral suppression of erythropoiesis in systemic lupus erythematosus (SLE) and rheumatoid arthritis.

Anemia due to inadequate red cell production often accompanies systemic lupus erythematosus and rheumatoid arthritis. We investigated its pathogenesis in 17 patients with these disorders, using a plasma clot culture system. In serum from normal donors and nonanemic patients CFU-E derived colony formation was not significantly altered by normal marrow cells (mean 74 +/- 12 colonies/6 x 10(4) cells), whereas colony formation was inhibited (mean 36 +/-6 colonies/6 x 10(4) cells) in serum from 10 anemic patients. In serum from anemic patients proliferation of the more primitive BFU-E was also reduced in three cases. In two patients with a humoral inhibitor, colony growth was suppressed by autologous marrow cells. In another patient without an inhibitor, colony formation was not suppressed by autologous bone marrow. The physical properties of this inhibitor are compatible with those of an immunoglobulin. Moreover, its presence is related to disease activity and it can be removed by successful therapy with either corticosteroids or plasma exchange. Circulating inhibitors of erythropoiesis may play an important role in causing severe anemia in patients with these rheumatic diseases.

Adolescent↗
PubMed1978

Detection of hemophilia A carriers by use of frozen plasma samples.

The efficacy of using promptly frozen plasma samples in the diagnosis of the carrier state for hemophilia A was evaluated by simultaneous measurement of factor VIII acitivity and antigen in 20 normal women and 20 obligate carriers. Factor VIII antigen was measured by two methods, electroimmunoassay and immunoradiometric assay. When the factor VIII activity and antigen data were evaluated by regression analysis, 94% of the carriers were correctly identified at the 95% confidence level.

Antigens↗