CRKI (SH2-SH3) and CRKII (SH2-SH3-SH3) are splicing isoforms of the oncoprotein CRK that regulate transcription and cytoskeletal reorganization for cell growth and motility by linking tyrosine kinases to small G proteins. CRKI shows substantial transforming activity, whereas the activity of CRKII is low, and phosphorylated CRKII has no biological activity whatsoever. The molecular mechanisms underlying the distinct biological activities of the CRK proteins remain elusive. We determined the solution structures of CRKI, CRKII and phosphorylated CRKII by NMR and identified the molecular mechanism that gives rise to their activities. Results from mutational analysis using rodent 3Y1 fibroblasts were consistent with those from the structural studies. Together, these data suggest that the linker region modulates the binding of CRKII to its targets, thus regulating cell growth and motility.

Secretin was injected into a feeding or nonfeeding artery of a gastrinoma and blood samples were taken from the hepatic vein (HV) or a peripheral artery (PA) to measure the changes of serum immunoreactive gastrin concentration (IRG). The IRG in the HV rose within 40 seconds and in the PA rose within 60 seconds after the injection of secretin into a feeding artery, but not after secretin was injected into a nonfeeder. These results indicated that secretin directly stimulates a gastrinoma to release gastrin in vivo. The selective arterial secretin injection test (SASI test) was applied in three patients in whom gastrinomas could not be located by computed tomography, ultrasonography, or arteriography, and functioning gastrinomas were located in all three patients. In one patient, malignant gastrinomas in the head of the pancreas and in the duodenum could be resected radically with the help of this test.S INCE ISENBERG ET AL.' discovered the paradoxical rise of serum gastrin levels after the intravenous injection ofsecretin in patients with the ZollingerEllison syndrome (ZE syndrome), the differential diagnosis of the ZE syndrome has become easier.2 In vitro studies of this reaction have shown that secretin directly stimulates gastrinoma to release gastrin.3'4 To ascertain whether this is also true in vivo, secretin was injected into the feeding arteries of the tumors visualized by arteriography in two patients with the ZE syndrome, and serial changes in the serum immunoreactive gastrin concentration (IRG) in the hepatic vein (HV)

Nonparenchymal cells, derived from transplanted BM, acquired HSC characteristics in both quiescent and activated states.

Germline chimeric chickens were produced by transfer of primordial germ cells from White Leghorn to Barred Plymouth Rock, and vice versa. Blood was collected from stage 13-15 embryos and primordial germ cells were concentrated by Ficoll density gradient centrifugation. Approximately 200 primordial germ cells were injected into the bloodstream through the dorsal aorta of stage 14-15 recipient embryos from which blood had been drawn via the dorsal aorta prior to the injection. Intact embryos were also prepared as recipients for White Leghorns only. The manipulated embryos were cultured in recipient eggshells until hatching. Germline chimerism of the chickens reaching maturity was examined by mating them with Barred Plymouth Rocks and donor-derived offspring were identified based on their feather color. The efficiency of production of germline chimeras was 95% (19/20). When primordial germ cells were transferred from White Leghorn to Barred Plymouth Rock, the average frequency of donor-derived offspring was 81% for three male chimeras (96% for one female chimera), and it was approximately 3.5 times higher for transfer in the opposite direction (23% for 6 male chimeras). Removing blood from recipient embryos prior to primordial germ cell injection enhanced the frequency of donor-derived offspring by 10% in resulting male chimeras. Male chimeras produced donor-derived offspring more frequently (approximately 3.8 times) than female chimeras. Increases, decreases, or no changes were observed in the frequency of donor-derived offspring from the germline chimeras with increasing age.(ABSTRACT TRUNCATED AT 250 WORDS)

No abstract

It is difficult to induce the maturation of embryonic stem (ES) cells into hepatocytes in vitro. We previously reported that Thy1-positive mesenchymal cells derived from the mouse fetal liver promote the maturation of hepatic progenitor cells. Here, we isolated alpha-fetoprotein (AFP)-producing cells from mouse ES cells for subsequent differentiation into hepatocytes in vitro by coculture with Thy1-positive cells. ES cells expressing green fluorescent protein (GFP) under the control of an AFP promoter were cultured under serum- and feeder layer-free culture conditions. The proportion of GFP-positive cells plateaued at 41.6 +/- 12.2% (means +/- SD) by day 7. GFP-positive cells, isolated by flow cytometry, were cultured in the presence or absence of Thy1-positive cells as a feeder layer. Isolated GFP-positive cells were stained for AFP, Foxa2, and albumin. The expression of mRNAs encoding tyrosine amino transferase, tryptophan 2,3-dioxygenase, and glucose-6-phosphatase were only detected following coculture with Thy1-positive cells. Following coculture with Thy1-positive cells, the isolated cells produced and stored glycogen. Ammonia clearance activity was also enhanced following coculture. Electron microscopic analysis indicated that the cocultured cells exhibited the morphologic features of mature hepatocytes. In conclusion, coculture with Thy1-positive cells in vitro induced the maturation of AFP-producing cells isolated from ES cell cultures into hepatocytes.

An experiment was carried out to investigate whether thick albumen is essential for the normal development of the chick embryo. Fertilized ova recovered from the oviducts of hens were cultured in vitro and transferred to recipient eggshells with (method A) or without (method B) replacement of the thick albumen by thin albumen. Embryos from freshly laid eggs were transferred to recipient eggshells with (method C) or without (method E) replacement of the thick albumen by thin albumen or with replacement of the thick albumen by thin albumen diluted with solution of salts (method D). Embryos were then incubated until hatching. The rates of hatching of the cultured embryos were 34.4%, 16.2%, 50.0%, 6.9-26.7%, and 47.5% for methods A, B, C, D, and E, respectively. Thus the rate of hatching of cultured fertilized ova was increased by replacement of the thick albumen by thin albumen at the blastoderm stage. Chicks obtained by method A reached maturity and produced viable offspring, and this technique provides an improved method for the culture of fertilized ova to hatching.

Previously, we reported a system to enrich mouse fetal hepatic progenitor cells (HPCs) by forming cell aggregates. In this study, we sorted two cell populations, CD49f ؉ Thy1 ؊ CD45 ؊ cells (