Various factors are known to regulate cell growth and differentiation, but less is known of agents which affect movement and positioning, particularly in epithelial-mesenchymal interactions. Cultured human embryo fibroblasts release a protein with a relative molecular mass (Mr) of approximately 50,000 (50K) that affects epithelial cells by causing a disruption of junctions, an increase in local motility and a scattering of contiguous sheets of cells. To investigate specificity, a range of cells has been examined for the ability to produce the factor and for sensitivity to its action. Most freshly isolated normal epithelia and epithelia from cell lines of normal tissue, but not epithelia from tumour cell lines or fibroblasts, were sensitive to scatter factor. In contrast, production of the factor, as identified by activity and by chromatography, was restricted to embryonic fibroblasts and certain variants of 3T3 and BHK21 cells and their transformed derivatives. We conclude that the scatter factor is a paracrine effector of epithelial-mesenchymal interaction, which affects the intercellular connections and mobility of normal epithelial cells. The factor might be involved in epithelial migration, such as occurs in embryogenesis or wound healing.

Synthetic oligonucleotides containing GC-rich triplet sequences were used in a scanning strategy to identify unstable genetic sequences at the myotonic dystrophy (DM) locus. A highly polymorphic GCT repeat was identified and found to be unstable, with an increased number of repeats occurring in DM patients. In the case of severe congenital DM, the paternal triplet allele was inherited unaltered while the maternal, DM-associated allele was unstable. These studies suggest that the mutational mechanism leading to DM is triplet amplification, similar to that occurring in the fragile X syndrome. The triplet repeat sequence is within a gene (to be referred to as myotonin-protein kinase), which has a sequence similar to protein kinases.

Scatter factor is a fibroblast-derived protein that causes separation of contiguous epithelial cells and increased local mobility of unanchored cells. Highly purified scatter factor has been obtained by a combination of ionexchange and reverse-phase chromatography from serum-free medium conditioned by a ras-transformed clone (D4) of mouse NIH 3T3 fibroblasts. Under nonreducing conditions scatter factor has a pI of -9.5 and migrates in SDS/polyacrylamide gels as a single band at -62 kDa from which epithelial scatter activity can be recovered. Treatment with reducing agents destroys biological activity and is associated with the appearance of two major bands at "57 and "30 kDa. Whether both the 57-kDa and 30-kDa polypeptides are required for biological activity remains to be established. All the activities observed in crude medium conditioned by cells producing scatter factor are retained by highly purified preparations of scatter factor.These include (i) increased local movement, modulation of morphology, and inhibition of junction formation by single epithelial cells and (it) disruption of epithelial interactions and cell scattering from preformed epithelial sheets. These changes occur with picomolar concentrations of purified scatter factor and without an effect on cell growth.Cell movement is restricted by the interaction with basement and extracellular membrane proteins (1) and, in certain tissues, by cell-cell interactions that involve cell-specific adhesion molecules (2). Movement of epithelial cells is further limited by cell-cell (desmosomes, tight and gap junctions) and cell-substratum (hemidesmosomes) junctional systems. Cytokines are also involved in the regulation of cell movement, as it appears that certain growth factors, including nerve growth factor (3), platelet-derived growth factor (4), and epidermal growth factor (5, 6) may stimulate cell movement as well as cell growth.There is increasing evidence, however, for a new group of cytokines that regulate cell movement with little or no effect on cell growth. Pioneering studies by Yoshida et al. (7) indicated that certain mouse and rat hepatoma lines and mouse and human leukemias produced a protein of 70 kDa that was chemotactic for the producer cells as well as for other tumor cells but not for polymorphonuclear cells (7). More recently, another motility factor has been isolated from serum-free medium conditioned by the human melanoma line A2058. This 55-kDa protein has both chemotactic and chemokinetic activity for the producer cells (but not for polymorphonuclear cells) and has been designated autocrine motility factor (AMF) (8). ras-transformed derivatives of mouse NIH 3T3 fibroblasts also produce AMF and respond to it and, interestingly, normal NIH 3T3 firboblasts, which do not produce AMF, are able to respond to the AMF secreted by ras-tranformed cells (8). A factor similar to AMF has been isolated from serum-free medium conditioned by a highly metastatic clone (MTLn3) of rat mammary adenocarcinoma (9). This factor (53 kDa) is chemota...

No abstract

Congenital heart defects comprise the most common form of major birth defects, affecting 0.7% of all newborn infants. Jacobsen syndrome (11q-) is a rare chromosomal disorder caused by deletions in distal 11q. We have previously determined that a wide spectrum of the most common congenital heart defects occur in 11q-, including an unprecedented high frequency of hypoplastic left heart syndrome (HLHS). We identified an approximately 7 Mb 'cardiac critical region' in distal 11q that contains a putative causative gene(s) for congenital heart disease. In this study, we utilized chromosomal microarray mapping to characterize three patients with 11q- and congenital heart defects that carry interstitial deletions overlapping the 7 Mb cardiac critical region. We propose that this 1.2 Mb region of overlap harbors a gene(s) that causes at least a subset of the congenital heart defects that occur in 11q-. We demonstrate that one gene in this region, ETS-1 (a member of the ETS family of transcription factors), is expressed in the endocardium and neural crest during early mouse heart development. Gene-targeted deletion of ETS-1 in mice in a C57/B6 background causes, with high penetrance, large membranous ventricular septal defects and a bifid cardiac apex, and less frequently a non-apex-forming left ventricle (one of the hallmarks of HLHS). Our results implicate an important role for the ETS-1 transcription factor in mammalian heart development and should provide important insights into some of the most common forms of congenital heart disease.

Comparison of mammalian cardiac ␣-and ␤-myosin heavy chain isoforms reveals 93% identity. To date, genetic methodologies have effected only minor switches in the mammalian cardiac myosin isoforms. Using cardiac-specific transgenesis, we have now obtained major myosin isoform shifts and/or replacements. Clusters of non-identical amino acids are found in functionally important regions, i.e. the surface loops 1 and 2, suggesting that these structures may regulate isoform-specific characteristics. Loop 1 alters filament sliding velocity, whereas Loop 2 modulates actin-activated ATPase rate in Dictyostelium myosin, but this remains untested in mammalian cardiac myosins. ␣ 3 ␤ isoform switches were engineered into mouse hearts via transgenesis. To assess the structural basis of isoform diversity, chimeric myosins in which the sequences of either Loop 1؉Loop 2 or Loop 2 of ␣-myosin were exchanged for those of ␤-myosin were expressed in vivo. 2-fold differences in filament sliding velocity and ATPase activity were found between the two isoforms. Filament sliding velocity of the Loop 1؉Loop 2 chimera and the ATPase activities of both loop chimeras were not significantly different compared with ␣-myosin. In mouse cardiac isoforms, myosin functionality does not depend on Loop 1 or Loop 2 sequences and must lie partially in other non-homologous residues.Myosin, the molecular motor of the heart, generates force and motion by coupling its ATPase activity to its cyclic interaction with actin. Myosin is a hexameric protein and is composed of two heavy chains (MHC) 1 and two essential and two regulatory myosin light chains. Structurally, MHC is composed of a number of discrete domains: a helical rod necessary for thick filament formation, and a globular head that contains the actin-binding site, catalytic, and motor domains (1).In the mammalian heart, two functionally distinct MHC isoforms, termed V 1 and V 3 , are present. V 1 is a homodimer of two ␣-MHC molecules, whereas V 3 is a ␤␤-homodimer. Expression of V 1 and V 3 is controlled both developmentally and hormonally. In the mouse, ␤-MHC expression in the ventricles predominates prenatally. However, via thyroid hormone regulation, ␤-MHC expression is silenced at birth, and ␣-MHC is transcribed (2). The functional differences between V 1 and V 3 myosin in terms of shortening velocity, force generation, and ATPase activity are profound. For example, rabbit V 1 myosin has a 2-3-fold faster actin filament sliding velocity than V 3 , but generates only half the average isometric force (3, 4). Likewise, both the Ca 2ϩ -stimulated and actin-activated ATPase activities of rabbit V 1 myosin are ϳ2-3 times greater than for V 3 myosin (3, 5). Similar differences in actin velocity and myofibrillar ATPase activity have been observed between mouse V 1

AT1 receptors are selectively downregulated in failing human ventricles, similar to the selective downregulation of beta 1 receptors. The relative lack of AT1 downregulation in ISC hearts may be related to differences in the degree of ventricular dysfunction.

We performed a multicenter, double-blind, randomized study to evaluate the effect of diltiazem on reinfarction after a non-Q-wave myocardial infarction. Nine centers enrolled 576 patients: 287 received diltiazem (90 mg every six hours) and 289 received placebo. Treatment was initiated 24 to 72 hours after the onset of infarction and continued for up to 14 days. The primary end point, reinfarction, was defined as an abnormal reelevation of MB creatine kinase in plasma within 14 days. Reinfarction occurred in 27 patients in the placebo group (9.3 percent) and in 15 in the diltiazem group (5.2 percent)--a 51.2 percent reduction in cumulative life-table incidence (P = 0.0297; 90 percent confidence interval, 7 to 67 percent). Diltiazem reduced the frequency of refractory postinfarction angina (a secondary end point) by 49.7 percent (P = 0.0345; 90 percent confidence interval, 6 to 73 percent). Mortality was similar in the two groups (3.1 and 3.8 percent, respectively, in the placebo and diltiazem groups), but adverse drug reactions (most of which were mild) were more common in the diltiazem group. Nevertheless, the drug was well tolerated, despite concurrent treatment with beta-blockers in 61 percent of the patients. We conclude that diltiazem was effective in preventing early reinfarction and severe angina after non-Q-wave infarction and that it was also safe and generally well tolerated.