Role of phosphatidylinositol-3 kinase and its association with Gab1 in thrombopoietin-mediated up-regulation of platelet function.

OBJECTIVE: Human blood platelets are easily available physiologic target cells for thrombopoietin (TPO). TPO up-regulates platelet aggregation and alpha-granule secretion induced by various agonists. We investigated the role of phosphatidylinositol 3-kinase (PI3K) and its association with Gab1 in TPO-mediated up-regulation of platelet function. MATERIALS AND METHODS: PI3K inhibitors (wortmannin and LY294002) and a MAP/ERK-kinase (MEK) inhibitor (PD98059) were used to investigate the role of these kinases in TPO-mediated up-regulation of platelet function. To elucidate the molecules associated with PI3K, we performed immunoprecipitation and pull-down experiments followed by immunoblotting. In vitro kinase assay also was performed to detect extracellular signal-regulated kinase (ERK) kinase activity. RESULTS: TPO up-regulated platelet alpha-granule secretion and aggregation induced by thrombin, which was dose-dependently inhibited by preincubation with wortmannin or LY294002. Immunoprecipitation and pull-down experiments revealed that regulatory subunit of PI3K, p85, was rapidly associated with tyrosine-phosphorylated Gab1 via its n- and c-terminal SH2 domains. Pretreatment of platelets with TPO dramatically augmented the thrombin-induced ERK activation, which was almost completely inhibited by LY294002. Furthermore, a MEK inhibitor, PD98059, not completely but significantly inhibited TPO-mediated up-regulation of thrombin-induced alpha-granule secretion. CONCLUSION: TPO induces the association of tyrosine-phosphorylated Gab1 with p85-PI3K. In downstream signaling, ERK is PI3K-dependently activated, which plays a critical role for TPO-mediated up-regulation of platelet function.

Failure to detect genetic alteration of the mannose-6-phosphate/insulin-like growth factor 2 receptor (M6P/IGF2R) gene in hepatocellular carcinomas in Japan.

The mannose-6-phosphate/insulin-like growth factor 2 receptor (M6P/IGF2R) suppresses cell growth through binding to the insulin-like growth factor 2 (IGF2) and latent complex of the transforming growth factor-beta (TGF-beta). Recently, it was reported in the United States that loss of heterozygosity (LOH) and mutations in exons 27, 28, and 31 of the M6P/IGF2R gene are frequent in hepatocellular carcinomas (HCCs) and adenomas. In view of the possible importance of this finding, especially for differential diagnosis of small hepatic lesions, we analyzed 43 primary HCCs, 2 adenomatous hyperplasias (AHs), and 3 regenerative nodules (RNs) developing in 42 Japanese patients in Japan for LOH using the polymorphic locus and for mutations by both single strand conformation polymorphism (SSCP) and direct sequencing methods. In the LOH study, 21 out of 22 informative HCCs and all of the informative AHs and RNs showed no allelic loss. In mutational studies of exons 27, 28, and 31, no mutations were detected either by SSCP or direct sequencing analysis in any of the 48 lesions. Thus inactivation of the M6P/IGF2R gene because of genetic alteration does not appear to be essential for hepatocarcinogenesis in Japan.

Novel mouse microsatellites: primer sequences and chromosomal location.

Sixty-nine sequences containing microsatellites were determined by analysis of clones from a pUC118 library of total genomic mouse DNA. These sequences were examined for size variation using polymerase chain reaction and gel electrophoresis. Fifty-one of them showed allelic variations between C57BL/6 and MSM, the two strains used for genetic mapping. Hence, their chromosomal location was determined using a panel consisting of 131 backcross mice that had been typed with 85 anchor loci. The microsatellites were distributed to most chromosomes except for chromosomes 16 and 19. These novel markers with defined locations are useful in linkage and genome mapping studies.

The influence of coronary artery bypass grafting to totally occluded coronary arteries on the left ventricular contractility.

Sixteen patients underwent 16 coronary artery bypass grafts (CABG) to totally occluded coronary arteries (TOCA), including 12 left anterior descending coronary arteries (LAD) and 4 right coronary arteries (RCA). Of these 16 CABGs, 2 of the RCA grafts became obstructed postoperatively and the remaining 14 patients with patent grafts were divided into the two following groups: 8 with previous infarcts in the region perfused by the TOCA (Group I) and 6 with no previous infarcts (Group II). The left ventricular (LV) ejection fraction and the mean verocity of circumferential fiber shortening significantly increased postoperatively in both groups and the PLVSP/LVESV significantly increased postoperatively in Group II. The LV segmental wall motion (SWM) in the region of TOCA significantly increased postoperatively in both groups. In 5 of the Group I patients, whose anterior and apical SWM was less than the lowest value of the normal subjects, the anterior and apical SWM significantly increased postoperatively. We thus concluded that CABG to totally occluded LAD results in an excellent graft patency rate, a significant improvement of SWM in the region of the TOCA and global LV contractility, even in patients with severe segmental dysfunction due to previous infarcts.

[A case report of aortic and mitral regurgitation combined with ankylosing spondylosis].

The purpose of this communication is to present a successful case of surgical treatment of aortic and mitral valve regurgitation associated with ankylosing spondylosis. A 48-year-old male was admitted with symptoms of palpitation and shortness of breath. His heart murmur had been pointed out for 4 months prior to this admission. His previous history did not show any evidence of rheumatic heart disease. Retrograde aortography showed moderate to severe aortic valvular regurgitation and echocardiography revealed mitral valve regurgitation with minor degree. At the same time he complained of rigidity of neck muscle as well as back pain for last 15 years. He underwent aortic valve replacement and mitral valve was left alone. His postoperative course was uneventful. In Japan, aortic valve regurgitation associated with ankylosing spondylosis has been reported to be very rare compared to the European or American people. Several important features in the diagnosis and operative treatment for this combination of diseases have been also discussed.