A nuclear isoform of myosin I beta that contains a unique 16-amino acid amino-terminal extension has been identified. An affinity-purified antibody to the 16-amino acid peptide demonstrated nuclear staining. Confocal and electron microscopy revealed that nuclear myosin I beta colocalized with RNA polymerase II in an alpha-amanitin- and actinomycin D-sensitive manner. The antibody coimmunoprecipitated RNA polymerase II and blocked in vitro RNA synthesis. This isoform of myosin I beta appears to be in a complex with RNA polymerase II and may affect transcription.

Orthotopic liver transplantation (LTx) is currently the only available treatment that has been proven to halt the progress of familial amyloidotic polyneuropathy (FAP). The aim of this study was to assess mortality and symptomatic response to LTx for FAP. All 86 FAP patients transplanted at our hospital between April 1990 and November 2005 were included in the study. Five patients underwent retransplantation. The 1-, 3-and 5-year patient survival rates in patients transplanted during 1996-2005 were 94.6%, 92.3% and 92.3%, respectively, a significant difference from the rates of 76.7%, 66.7% and 66.7%, respectively, during 1990-1995 (p = 0.0003). Multivariate analysis revealed that the age at the time of LTx (≥40 years), duration of the disease (≥7 years) and modified body mass index (mBMI) (<600) were independent prognostic factors for patient survival. A halt in the progress of symptoms was noted in most patients, but only a minority experienced an improvement after LTx. To optimize the posttransplant prognosis, LTx should be performed in the early stages of the disease, and close post-LTx monitoring of heart function by echocardiography and of heart arrhythmia by Holter ECG is mandatory.

To explore the role of nonmuscle myosin II isoforms during mouse gametogenesis, fertilization, and early development, localization and microinjection studies were performed using monospecific antibodies to myosin IIA and IIB isotypes. Each myosin II antibody recognizes a 205-kDa protein in oocytes, but not mature sperm. Myosin IIA and IIB demonstrate differential expression during meiotic maturation and following fertilization: only the IIA isoform detects metaphase spindles or accumulates in the mitotic cleavage furrow. In the unfertilized oocyte, both myosin isoforms are polarized in the cortex directly overlying the metaphase-arrested second meiotic spindle. Cortical polarization is altered after spindle disassembly with Colcemid: the scattered meiotic chromosomes initiate myosin IIA and microfilament assemble in the vicinity of each chromosome mass. During sperm incorporation, both myosin II isotypes concentrate in the second polar body cleavage furrow and the sperm incorporation cone. In functional experiments, the microinjection of myosin IIA antibody disrupts meiotic maturation to metaphase II arrest, probably through depletion of spindle-associated myosin IIA protein and antibody binding to chromosome surfaces. Conversely, the microinjection of myosin IIB antibody blocks microfilament-directed chromosome scattering in Colcemid-treated mature oocytes, suggesting a role in mediating chromosome-cortical actomyosin interactions. Neither myosin II antibody, alone or coinjected, blocks second polar body formation, in vitro fertilization, or cytokinesis. Finally, microinjection of a nonphosphorylatable 20-kDa regulatory myosin light chain specifically blocks sperm incorporation cone disassembly and impedes cell cycle progression, suggesting that interference with myosin II phosphorylation influences fertilization. Thus, conventional myosins break cortical symmetry in oocytes by participating in eccentric meiotic spindle positioning, sperm incorporation cone dynamics, and cytokinesis. Although murine sperm do not express myosin II, different myosin II isotypes may have distinct roles during early embryonic development. INTRODUCTIONThe cortex of mature mouse oocytes is polarized: the area adjacent to the eccentrically positioned second meiotic spindle is devoid of cortical granules and surface microvilli, diminished in concanavalin A lectin binding and demonstrates an increase in cortical actin filaments (reviewed by Longo, 1989). Similar events are observed in the cortex and plasma membrane in the vicinity of the incorporating sperm head (Nicosia et al., 1977(Nicosia et al., , 1978Shalgi et al., 1978). The induction of these cortical and cell surface modifications are ‡ Corresponding author: Oregon Regional Primate Research Center and Oregon Health Science University, 505 N.W. 185th Avenue, Beaverton, OR 97006-3499. E-mail: schatten@ohsu.edu.© 1998 by The American Society for Cell Biology 2509 strongly correlated to the presence of meiotic chromosomes or demembranated sperm DNA (Longo and Chen, 1985;Maro et al.,...

Orthodox etching of HVPE-grown GaN in molten eutectic of KOH + NaOH (E etch) and in hot sulfuric and phosphoric acids (HH etch) is discussed in detail. Three size grades of pits are formed by the preferential E etching at the outcrops of threading dislocations on the Ga-polar surface of GaN. Using transmission electron microscopy (TEM) as the calibration tool it is shown that the largest pits are formed on screw, intermediate on mixed and the smallest on edge dislocations. This sequence of size does not follow the sequence of the Burgers values (and thus the magnitude of the elastic energy) of corresponding dislocations. This discrepancy is explained taking into account the effect of decoration of dislocations, the degree of which is expected to be different depending on the lattice deformation around the dislocations, i.e. on the edge component of the Burgers vector. It is argued that the large scatter of optimal etching temperatures required for revealing all three types of dislocations in HVPE-grown samples from different sources also depends upon the energetic status of dislocations. The role of kinetics for reliability of etching in both etches is discussed and the way of optimization of the etching parameters is shown.

We produced and affinity-purified polyclonal antibodies to adrenal myosin I. These antibodies recognize adrenal myosin I by Western blot analysis (116 kDa) and inhibit the actin-activated ATPase activity of purified adrenal myosin I. They also recognize a 120-kDa protein in extracts prepared from many different cell lines. Fluorescence microscopy demonstrated the presence of immunoreactive material in the perinuclear region, the leading edges, and the nuclei of 3T3 cells. Fluorescence microscopy also demonstrated nuclear staining in mouse oocytes at the germinal vesicle stage and in the pronuclei during fertilization. Confocal and immunoelectron microscopy confirmed the intranuclear localization. Electron microscopy also demonstrated staining of structures in nucleoli that are thought to be associated with rDNA transcription. Western blot analyses revealed the presence of the 120-kDa protein in extracts prepared from nuclei that are apparently free of cytosolic contamination. The same nuclear protein binds 125 I-calmodulin and is photoaffinity labeled with [␣-32 P]ATP. The 120-kDa protein was partially purified from twice washed nuclei using ammonium sulfate fractionation and gel filtration chromatography. Column fractions containing 120-kDa protein as revealed by Western blot analysis also contain K ؉ -EDTA ATPase activity. The 120-kDa protein was also shown to bind actin in the absence, but not the presence, of ATP. Since K ؉ -EDTA ATPase activity, actin, and ATP binding are defining features of the members of the myosin superfamily of proteins, we propose that the 120-kDa protein is a previously undescribed myosin I isoform that is an intranuclear actin-based molecular motor.Myosin I is a single-headed, monomeric, actin-activated ATPase (1). First described in Acanthamoeba castellanii (1), myosin I is now known to be widely distributed in metazoan cells (1-12). As additional myosin I proteins have been identified, it has become clear that there are at least four different subclasses of myosin I (1). All myosin I proteins consist of a 110 -150-kDa heavy chain and 1-6 light chains located in the neck region between the head and tail (1). This light chain has been shown to be calmodulin in vertebrate myosin I proteins (1). Immunofluorescence studies of mammalian cells have shown that myosin I is diffusely distributed throughout the entire cytoplasm and that it concentrates near cortical surfaces and in the perinuclear region (12, 13). Although evidence for specific roles of myosin I proteins in metazoan cells is lacking, there is speculation, based on localization studies, that myosin I proteins are molecular motors involved in plasma membrane extension (12, 13), vesicle and organelle transport (14), and mechanochemical regulation of calcium channels in hair cells (15). To investigate the role of myosin I in mammalian cells, we produced and affinity-purified polyclonal antibodies to adrenal myosin I. These antibodies recognize a 120-kDa protein that is found in the cytoplasm and the nucleus. Moreover, we present...

Microdialysis provides the opportunity to continuously monitor metabolic changes in tissue. The aim of the study is to monitor metabolic changes in the liver graft over time during transplantation in a pig model. Fourteen littermate female pigs with a body weight of 30 to 34 kg were used for seven orthotopic liver transplantations. Intrahepatic implantation of a microdialysis catheter into the liver graft was performed in the donor. Microdialysis samples were collected at 20-minute intervals during the donor operation, cold preservation, and for 7 hours after reperfusion in the recipient. Glucose, lactate, pyruvate, and glycerol concentrations were measured. After cold perfusion, glucose, lactate, and glycerol levels increased, whereas pyruvate levels decreased rapidly. During cold storage, glucose and glycerol levels increased, whereas lactate levels remained stable and pyruvate levels were undetectable. During implantation of the liver graft, glucose, lactate, and glycerol levels showed an accelerated increase. After portal reperfusion, glucose, lactate, and glycerol levels continued to increase for another 40 to 60 minutes, after which they decreased and finally settled at normal levels. At this time, pyruvate levels increased, with a peak within 2 hours after reperfusion, and then decreased to normal levels. Calculated lactate-pyruvate ratio increased after cold perfusion and remained stable during cold storage. During rewarming, it showed an accelerated increase, but after reperfusion, it decreased rapidly. Rewarming and reperfusion are most harmful to the liver, reflected by an accelerated increase in glucose and glycerol levels and lactate-pyruvate ratio. High intrahepatic glucose levels during ischemia appear to be a liver-specific event, which may represent glycogen degradation in injured hepatocytes. (Liver Transpl 2002;8:424-432.)

The monocyte population in blood is considered a possible source of endothelial precursors. Because endothelial-specific receptor tyrosine kinases act as regulators of endothelial cell function, we investigated whether expression of the vascular endothelial growth factor receptor-2 (VEGFR-2) on monocytes is important for their endothelial-like functional capacity. Peripheral-blood monocytes expressing vascular endothelial growth factor receptor-2 (VEGFR-2), or CD14 ؉ /VEGFR-2 ؉ , were isolated, and their phenotypic, morphologic, and functional capacities were compared with those of monocytes nega-

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