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D Coles

Publications and source records attributed to D Coles.

8 recordsLinked to original sources
PubMed1999

Using molecular genetics as a tool in understanding crawling cell locomotion in myoblasts.

We have used digitally recorded interference microscopy with automatic phase shifting (DRIMAPS) to investigate the crawling locomotion of normal and mutant mouse myoblasts. Contraction forces that give rise to cell body movement, tail retraction and cell adhesion to the substrate in myoblasts and other locomoting tissue cells arise from the interactions of actin and non-muscle myosin II. The activity of non-muscle myosin II is regulated differently from that of skeletal myosin. Using DRIMAPS, we found that crawling locomotion was altered in myoblasts that heterologously expressed human beta-cardiac myosin heavy chain (MHC); the cells moved more slowly and had reduced rates of protrusion and retraction. Immunolocalization demonstrated that MHC and non-muscle myosin II were not co-localized, suggesting that MHC does not compete directly with myosin II, but interferes with cell locomotion by binding inappropriately to actin filaments and possibly cross-linking them. Myosin I may be involved in protrusion of the lamellipodia. However, using DRIMAPS, we found that crawling locomotion was unaltered in myoblasts that heterologously expressed a truncated myosin I which lacked the membrane-binding tail domain. This suggests that, if endogenous myosin I is important for cell locomotion, this mutant was unable to interfere with its action. We conclude that the effects on locomotion of expressing foreign or mutant proteins of the cytoskeleton in vertebrate cells can be subtle and can be swamped by the intrinsic variability of the cells. Their characterization requires automated methods of acquiring data, such as DRIMAPS, and careful statistical analysis in order to take account of other sources of variation.

Actins↗
PubMed1997

Preferential adhesion to and survival on patterned laminin organizes myogenesis in vitro.

We have examined a potential role for differential adhesiveness in muscle development using an in vitro model which employed the culture of myoblasts and myotubes, (conditionally immortal myogenic cells, H2k(b)-tsA58), on micropatterned surfaces. These surfaces are made up of multiple alternating tracks of hydrophobic organosilane-treated glass and untreated glass (track width ranging from 5 to 100 microm). We found that myoblasts were aligned on patterns in the presence of serum, by adhering to the tracks of untreated glass, which had preferentially adsorbed serum attachment factors. However, as serum attachment factors are not sufficient for maintenance of adhesion of mature myotubes, we determined whether precoating patterns with laminin, which maintains adhesion, could still provide a differential adhesive cue. Laminin preferentially adsorbs to the hydrophobic regions resulting in alternating tracks that have adsorbed laminin or serum attachment factors. Myoblasts were less well aligned on these patterns as they could adhere both to the untreated glass and to laminin on the previously hydrophobic tracks, but did show a preference for laminin. However, cell alignment increased upon differentiation into myotubes and continued to increase as the myotubes matured. We found that the alignment of myoblasts and myotubes on patterns increased as track width increased. In addition, adhesion to laminin was required for long term survival of the myotubes. Myotubes that had formed on nonlaminin surfaces began to detach after 2 days of differentiation. Although we found that myoblasts preferentially clustered on laminin tracks, this arrangement did not influence the diameter of the myotubes formed, upon differentiation. Instead, the number of myotubes per track increased with track width, while the myotube diameter remained constant. This uniformity of myotube diameter suggests that a mechanism exists which restricts the ability of myoblasts to undergo lateral fusion. Overall, these findings suggest that differential adhesiveness could be an important mechanism for formation and survival of myotubes, and by using these patterns we have demonstrated a mechanism controlling the formation of linear myotubes by restricting the geometry of cell-cell adhesion.

Animals↗
PubMed1997

Myogenic cells express multiple myosin isoforms.

In vivo and in vitro, proliferating motile myoblasts form aligned groups of cells, with a characteristic bipolar morphology, subsequently become post-mitotic, begin to express skeletal myosin and fuse. We were interested in whether members of the myosin superfamily were involved in myogenesis. We found that the myoblasts expressed multiple myosin isoforms, from at least five different classes of the myosin superfamily (classes I, II, V, VII and IX), using RT-PCR and degenerate primers to conserved regions of myosin. All of these myosin isoforms were expressed most highly in myoblasts and their expression decreased as they differentiated into mature myotubes, by RNAse protection assays, and Western analysis. However, only myosin I alpha, non-muscle myosin IIA and IIB together with actin relocalize in response to the differentiative state of the cell. In single cells, myosin I alpha was found at the leading edge, in rear microspikes and had a punctate cytoplasmic staining, and non-muscle myosin was associated with actin bundles as previously described for fibroblasts. In aligned groups of cells, all these proteins were found at the plasma membrane. Co-staining for skeletal myosin II, and myosin I alpha showed that myosin I alpha also appeared to be expressed at higher levels in post-mitotic myoblasts that had begun to express skeletal myosin prior to fusion. In early myotubes, actin and non-muscle myosin IIA and IIB remained localized at the membrane. All of the other myosin isoforms we looked at, myosin V, myosin IX and a second isoform of myosin I (mouse homologue to myr2) showed a punctate cytoplasmic staining which did not change as the myoblasts differentiated. In conclusion, although we found that myoblasts express many different isoforms of the myosin superfamily, only myosin I alpha, non-muscle myosin IIA and IIB appear to play any direct role in myogenesis.

Amino Acid Sequence↗
PubMed1997

Teachers' ratings of nursery class children's behaviour before and after availability of television by satellite.

Teachers rated children's behaviour in two nursery class cohorts on St. Helena, South Atlantic Ocean. Ratings were obtained for the 1993 cohort (n = 54) prior to the availability of satellite television and the 1995 cohort (n = 57) after the introduction of television. Analysis showed some reductions in ratings of children's adaptive behaviour after the availability of television.

Atlantic Islands↗
PubMed1978

Salivary neutralizing activity against herpes simplex virus type 1.

Samples of saliva collected from 40 patients, of whom 16 had a previous clinical history of herpes simplex virus type 1 (HSV-1) infection, were concentrated and sterilized. Salivary neutralizing activity, as measured by a plaque-reduction method, was found in 62% of those patients with a history of oral lesions. Studies with antisera to human immunoglobulin indicated that the IgG class of immunoglobulin was a major HSV-1-neutralizing component of both saliva and serum.

Adult↗