The Ca(2+) /calmodulin-dependent kinase kinase ß-AMP-activated protein kinase-α1 pathway regulates phosphorylation of cytoskeletal targets in thrombin-stimulated human platelets.

BACKGROUND: Platelet activation requires sweeping morphologic changes, supported by contraction and remodeling of the platelet actin cytoskeleton. In various other cell types, AMP-activated protein kinase (AMPK) controls the phosphorylation state of cytoskeletal targets. OBJECTIVE: To determine whether AMPK is activated during platelet aggregation and contributes to the control of cytoskeletal targets. RESULTS: We found that AMPK-α1 was mainly activated by thrombin, and not by other platelet agonists, in purified human platelets. Thrombin activated AMPK-α1 ex vivo via a Ca(2+) /calmodulin-dependent kinase kinase ß (CaMKKß)-dependent pathway. Pharmacologic inhibition of CaMKKß blocked thrombin-induced platelet aggregation and counteracted thrombin-induced phosphorylation of several cytoskeletal proteins, namely, regulatory myosin light chains (MLCs), cofilin, and vasodilator-stimulated phosphoprotein (VASP), three key elements involved in actin cytoskeletal contraction and polymerization. Platelets isolated from mice lacking AMPK-α1 showed reduced aggregation in response to thrombin, and this was associated with defects in MLC, cofilin and VASP phosphorylation and actin polymerization. More importantly, we show, for the first time, that the AMPK pathway is activated in platelets of patients undergoing major cardiac surgery, in a heparin-sensitive manner. CONCLUSION: AMPK-α1 is activated by thrombin in human platelets. It controls the phosphorylation of key cytoskeletal targets and actin cytoskeletal remodeling during platelet aggregation.

Novel tumor-promoting property of tamoxifen.

The tumor-promoting phorbol ester TPA (12-O-tetradecanoylphorbol-13-acetate) cooperates with c-Src overexpression to transform rat fibroblasts. TPA transforms c-Src-overexpressing cells by depleting the delta isoform of protein kinase C (PKCdelta). Tamoxifen, which has both estrogen-mimetic and estrogen-antagonist properties, has been widely used to improve the prognosis of breast cancer patients. However, with extended use, there is an increased risk for endometrial and other cancers that can be observed within 10 years of treatment. We report here that tamoxifen, similar to TPA, cooperates with c-Src overexpression to transform 3Y1 rat fibroblasts. Tamoxifen induced both DNA synthesis and anchorage-independent cell proliferation in c-Src-overexpressing, but not in parental, 3Y1 rat fibroblasts. Tamoxifen also induced an association between c-Src and PKCdelta that resulted in the tyrosine phosphorylation and down-regulation of PKCdelta. These phenotypes were not induced by estrogen, indicating that the effect of tamoxifen was in addition to any estrogen-mimetic effects. Thus, in addition to the hyperplasia-inducing capability of an estrogen-mimetic, tamoxifen has an additional tumor-promoting capability similar to that of TPA. The dual tumor-promoting capability of both estrogen- and TPA-mimetic properties for tamoxifen may contribute to the increased incidence of endometrial cancers observed in the relatively short exposure period of <10 years.

Association between v-Src and protein kinase C delta in v-Src-transformed fibroblasts.

In response to the kinase activity of v-Src there is an increase in the membrane association of the novel protein kinase C (PKC) isoform PKC delta (Zang, Q., Frankel, P., and Foster, D. A. (1995) Cell Growth Differ. 6, 1367-1373). We report here that in v-Src-transformed cells PKC delta co-immunoprecipitates with v-Src and is phosphorylated on tyrosine. The tyrosine-phosphorylated PKC delta had reduced enzymatic activity relative to the non-tyrosine-phosphorylated PKC delta from v-Src-transformed cells. The association between Src and PKC delta was dependent upon both an active Src kinase and membrane association. The association between c-Src Y527F and PKC delta was substantially enhanced by mutating a PKC phosphorylation site at Ser-12 in Src to Ala indicating that PKC delta phosphorylation of Src at Ser-12 destabilizes the interaction, possibly in a negative feedback loop. These data demonstrate that upon recruitment of PKC delta to the membrane in v-Src-transformed cells there is the formation of a Src.PKC delta complex in which PKC delta becomes phosphorylated on tyrosine and down-regulated.

[9 cases of early gastric cancer disclosed on chronic ulcer with or without complications]. / Considérations sur 9 cas de early gastric cancer découverts sur ulcère peptique chronique, compliqué ou non.

The authors report their experience on 9 cases of early gastric cancer developed on a peptic ulcer border. They outline the paradoxical providence of the complications (hemorrhage and perforation) leading to cancer recognition as a focus in the benign ulcer. Therapeutic management of the occasionally discovered cancer on the peptic ulcer border is also discussed.