The SH2-containing adapter protein GRB10 interacts with BCR-ABL.

Bcr-Abl is an oncogenic tyrosine kinase expressed in tumor cells of CML and a subset of ALL which in its unregulated and activated state is thought to cause cell transformation and leukemia. Bcr-Abl contains several autophosphorylation sites which serve as potential docking sites for SH2-containing signaling molecules. Mutational analysis has indicated that these autophosphorylation sites play a critical role in the transforming capability of Bcr-Abl. It has been shown that the SH2-containing adapter protein Grb2 binds to the autophosphorylation site Tyr(p)177 whereby it couples Bcr-Abl to the Ras pathway. The biological consequences of this interaction, however, are presently unclear. A Tyr177-mutated Bcr-Abl which lacks the ability to interact with the Grb2-SH2 domain still transforms myeloid cells and generates tumors in nude mice. We performed a yeast two-hybrid screen to identify signaling proteins which bind to distinct Bcr-Abl autophosphorylation sites. Autophosphorylation of Bcr-Abl in yeast was accomplished by using the DNA binding protein LexA which permits dimerization and crossphosphorylation of the fused bait. Using a LexA-Bcr-Abl full length fusion protein as bait, we identified several SH2-containing proteins. Among them we confirmed molecules already shown by others to interact with Bcr-Abl, in vivo, including Grb2, PI-3-kinase and Crk indicating that dimerization in yeast leads to autophosphorylation of tyrosine residues crucial for Bcr-Abl signaling in vivo. More importantly, we identified the SH2-containing protein Grb10 as a new binding partner for Bcr-Abl. This binding occurs in a phosphotyrosine-dependent manner at Bcr sites of Bcr-Abl. Both Abl and Bcr alone, as well as a kinase-defective Bcr-Abl, failed to interact with Grb10 in yeast. Mutational analysis uncovered a new SH2 binding site in Bcr-Abl located between Bcr aa242-446, which is different from the Grb2 binding site. Binding could be demonstrated in vitro and also in vivo as shown by co-immunoprecipitation analysis in CML cells. Using a temperature sensitive Bcr-Abl stably overexpressed in hematopoetic cells, we demonstrated that complex formation of Grb10 with Bcr-Abl was kinase activation-dependent in vivo. Notably, a Bcr-Abl mutant protein (Bcr/1-242-Abl) which lacks the ability to interact with Grb10 partially alleviated IL-3 dependence of Ba/F3 cells, indicating that the Grb10/Bcr-Abl interaction is important for Bcr-Abl-induced IL-3 independence of Ba/F3 cells. In addition, the Bcr/1-242-Abl mutant has a reduced capacity to induce focus formation in fibroblasts.

Autonomic manipulation influences both temporal and frequency analyses of late potentials.

Previous studies of late potentials have not standardized the autonomic milieu at the time of testing. We studied the effects of autonomic manipulation in seven patients with previous Q wave myocardial infarction. Late potentials were evaluated using standard temporal (TD) and spectral temporal mapping techniques (STM) in the drug free state, and during separate intravenous administration of each of the following: isoproterenol, esmolol, and atropine. Isoproterenol was titrated to achieve a heart rate of 130% of baseline. Esmolol was infused at a rate of 250 micrograms/kg per minute, after a loading dose of 500 micrograms/kg. Atropine was given as a 2-mg bolus. In addition, five patients who received no drug infusions acted as controls, undergoing four serial signal-averaging studies in the baseline state: a "baseline" study, and then three additional studies at time intervals similar to those incurred by the study patients. Therefore, a total of 21 TD and 21 STM tests were done in the study group (seven patients; three drugs per patient) during the drug infusions, and 15 TD and 15 STM tests were done in the control group (five patients; three "nonbaseline" tests per patient). A change (normal to abnormal, or vice versa) in TD during a drug infusion occurred in 24% of the tests. No such change occurred in the control group (P < 0.01). A change in STM during a drug infusion occurred in 38% of tests, versus 13% of tests in the control group (P = 0.14). Overall, six of seven patients had a change in TD and/or STM diagnosis with infusion of one or more of the study drugs.(ABSTRACT TRUNCATED AT 250 WORDS)

Transesophageal echocardiography in the diagnosis of ostial left coronary artery stenosis.

The diagnosis of ostial stenosis of the left main coronary artery is usually made by use of coronary angiography. However, positioning of the catheter across the obstruction may obscure this diagnosis during contrast injection. Although a damping of arterial pressure when the catheter enters the left coronary artery may suggest ostial stenosis, it may not be possible to make this diagnosis with certainty during cardiac catheterization. We report a series of four patients in whom the left coronary ostium and proximal left coronary arteries were visualized by means of transesophageal echocardiography. Both ostial narrowing by plaque and abnormally fast flow velocities were seen. In each case the echocardiographic findings contributed to the subsequent management of the patients.

Metastatic cardiac liposarcoma: diagnosis by transesophageal echocardiography and magnetic resonance imaging.

The most superior portion of the right atrium is not well visualized by transthoracic echocardiography. This limits the ability of the technique to detect intracardiac disease in this area. We describe a 41-year-old man with a history of liposarcoma in whom transthoracic echocardiography was unable to elucidate a right atrial metastasis. Transesophageal echocardiography demonstrated the morphology and extent of the large right atrial mass. These findings were well correlated with both magnetic resonance imaging and surgery.

Pitfalls in the diagnosis of thoracic aortic aneurysm by transesophageal echocardiography.

A 74-year-old man underwent transesophageal echocardiography to evaluate the possibility of dissection of the descending aorta. The study demonstrated a round lumen that contained an echogenic mass mimicking aortic aneurysm with thrombus. However, computerized tomographic scanning identified the lumen as the pleural cavity containing a collapsed lung. The esophagus was on the right side of the spine, and the transesophageal technique could not visualize the descending aorta.

Pneumomediastinum complicating Pneumocystis carinii pneumonia in a patient with AIDS.

The destructive potential of Pneumocystis carinii infection in patients with AIDS has been amply documented. This report describes a homosexual man with P. carinii pneumonia (PCP) complicated by pneumatocoeles and pneumomediastinum. The pneumomediastinum almost completely resolved with successful antimicrobial therapy. Pneumomediastinum, like pneumothorax, should be anticipated in patients with AIDS and PCP.

Echocardiographic diagnosis of bland thrombus on an aortic valve.

Echocardiography is a valuable modality for imaging thrombi and vegetations. Although patients with systemic lupus erythematosus may develop Libman-Sacks endocarditis, in which punctate verrucous plaques are found histologically on valvular endocardial tissue, these lesions are ordinarily too small for ultrasonic imaging. We describe a patient with systemic lupus erythematosus complicated by systemic embolization, who had a large mobile aortic valve thrombus, which was discovered with the aid of echocardiography and was confirmed by surgical resection.

Use of sequential cardiac enzyme analysis in stratification of risk for myocardial infarction in patients with unstable angina.

Patients presenting to the emergency room with unstable angina are a challenge to physicians whose responsibility it is to ration access to coronary care unit beds, a resource that is often in short supply. In this study, initial cardiac enzyme analysis was not helpful in identifying patients having an acute myocardial infarction. However, using two enzyme determinations, performed at least six hours apart, we were able to identify a large percentage of our patients with unstable angina who were at low risk for myocardial infarction. Patients with two consecutive normal creatinine kinase levels had only a 2% chance of having an acute myocardial infarction. We believe these data can assist physicians where monitored beds are in short supply to make judicious use of this limited resource; the subsequent reduction in critical care unit admissions and duration of stay could result in substantial monetary savings.