A national survey of endoscopic mucosal resection for superficial gastrointestinal neoplasia.

BACKGROUND AND STUDY AIM: Studies on endoscopic mucosal resection (EMR) are mostly based on data from centers with high expertise. We report the average EMR results in a national survey of consecutive patients in France. METHODS: A 1-year survey was carried out to record immediate outcome data of all EMRs performed, regardless of lesion size or gastrointestinal location. RESULTS: Overall, 1335 EMRs in 1210 patients were reported by 241 of the 736 gastroenterologists who performed such procedures (33 %). Resections were done for upper gastrointestinal lesions in 125 cases (41 esophageal, 43 gastric, and 41 duodenal lesions), in 45 % of cases using specific EMR techniques such as ligation, cap, or traction. The technique for resecting the 1210 lower gastrointestinal lesions mostly consisted of saline-assisted polypectomy or EMR, with specific techniques used in only 2.2 %. En bloc resection was less common with esophageal (46 %) or duodenal (54 %) neoplasms than in the lower gastrointestinal tract (73 %); size also had some influence (53 % > 1 cm vs. 92 % ≤ 1 cm). The overall complication rate was 5.2 %; the rate was lower for lesions 1 cm or smaller (0.6 % vs. 4.6 %). Fifty-four early and 17 delayed complications were recorded, in 12 % of upper gastrointestinal and 4.6 % of colonic lesions. Surgery became necessary in 1.6 % for upper and 2.9 % for lower gastrointestinal neoplasms. No association was seen between physician EMR caseload and either en bloc resection rate or complication rate. CONCLUSIONS: EMR in general, especially saline-assisted polypectomy in the colon, appears to be reasonably safe even when performed by nonexperts. EMR for larger or for upper gastrointestinal lesions should probably be limited to high-volume centers.

Evaluation of the Clot Signature Analyzer as a hemostasis test in healthy volunteers exposed to low doses of aspirin.

Several variables affect bleeding time that make it difficult to obtain consistent measurements. The Clot Signature Analyzer (CSA) has been developed to assess in vitro hemostasis using well-controlled flow chambers. In this study, the equivalencies in the CSA parameters with the conventional bleeding time or platelet aggregation methods were evaluated in subjects exposed to aspirin. The CSA parameters, platelet hemostasis time (PHT) and collagen-induced thrombus formation (CITF), were compared to bleeding time (Surgicutt2) and collagen-induced platelet aggregation, respectively. Fifty-three healthy volunteers were given two doses of aspirin (81 and 243 mg) in one day. Following the baseline period, the volunteers took 81 mg of aspirin and then took 243 mg 2 hours later. The changes in each value from the baseline to that at either aspirin dose (2 hours after dosing) were evaluated. Platelet hemostasis time and CITF correlated well with bleeding time and aggregation, respectively, but PHT was not significantly increased after 81 mg of aspirin, whereas bleeding time was significantly increased. The variation in PHT was slightly higher than that of bleeding time. At 81 mg, CITF was significantly increased but aggregation was not, even though the variation was comparable. This suggests that PHT and CITF can simulate the changes in bleeding time and aggregation, respectively, but the sensitivity of PHT for detecting the changes in bleeding time was no better than the conventional method. Also, CITF was more sensitive than aggregation in detecting platelet response to collagen. In conclusion, the proposed CSA is not always suitable for detecting hemostatic abnormalities.

Contributions of glycoprotein Ib and the seven transmembrane domain receptor to increases in platelet cytoplasmic [Ca2+] induced by alpha-thrombin.

The individual contributions of glycoprotein Ib (GPIb) and the seven transmembrane domain receptor (STDR) to increases in platelet [Ca2+]i induced by alpha-thrombin or the tethered ligand peptide (TLP; SFLLRNPNDKYEPF) have been determined in control platelets, in platelets where the thrombin binding site on GPIb was blocked with the monoclonal antibodies TM60 and LJ-Ib10, in platelets where access of thrombin to the STDR was blocked by polyclonal antipeptide antibodies, and in Bernard-Soulier platelets which constitutively lack GPIb. Curve-fitting analyses (LIGAND) showed that binding of PPACK-thrombin and alpha-thrombin to the moderate-affinity site was not detected in the best-fit model in the presence of anti-STDR antibodies although with alpha-thrombin there was also decreased binding at the high-affinity site. Conversely, TM60 blocked binding of alpha-thrombin to the high-affinity site but also decreased binding at the moderate affinity site. Separately, either TM60 or anti-TNA (150 micrograms/mL) reduced thrombin (0.5 nM)-induced elevations in [Ca2+]i to 50% of control values, but Ca2+ elevations were essentially abrogated (4.2 +/- 5%) when the two were added in combination. [Ca2+]i dose-response curves for alpha-thrombin were curvilinear and were only 50% of controls in the presence of anti-GPIb or anti-STDR antibodies at up to 10 nM alpha-thrombin, with their greatest sensitivity being below 2 nM. With Bernard-Soulier platelets, changes in [Ca2+]i were not detectable at < or = 0.5 nM alpha-thrombin but were also 50% of controls at 5-10 nM alpha-thrombin. [Ca2+]i responses to TLP (1-100 microM) of antibody-blocked platelets were identical to those of controls whereas responses were approximately 50% of controls in Bernard-Soulier platelets. The rate of increase in [Ca2+]i in controls was twice that seen in antibody-blocked platelets and about 5-fold greater than in Bernard-Soulier platelets. These results demonstrate that both GPIb and the STDR are required to ensure the optimal rate and extent of platelet activation over a range of alpha-thrombin concentrations (0.3-10 nM) and that the STDR corresponds to the previously described moderate-affinity thrombin receptor.

Differentiation of the two forms of GPIb functioning as receptors for alpha-thrombin and von Willebrand factor: Ca2+ responses of protease-treated human platelets activated with alpha-thrombin and the tethered ligand peptide.

Previous results have shown that both GPIb and the seven transmembrane domain receptor (STDR) are required for optimal thrombin-induced platelet activation (Greco et al., 1996). Limited degradation (approximately 10%) of GPIb and the STDR by elastase reduced the Ca2+ response to 0.5 nM alpha-thrombin by only 10% whereas Serratia marcescens metalloprotease reduced the Ca2+ response by 80% and fully abrogated high-affinity thrombin binding and aggregation. vWF/ristocetin-induced agglutination was only slightly reduced (20%) while Ca2+ and aggregation response to higher thrombin concentrations were retained. At increasing elastase and Serratia protease concentrations, degradation of the STDR proceeded from the amino-terminal domain, but Ca2+ responses to the tethered ligand peptide SFLLRNPNDKYEPF were not affected by either protease. These results show that both putative thrombin receptors are susceptible to protease degradation and suggest that Serratia protease is able to differentiate the GPIb-mediated events associated with thrombin activation from those associated with ristocetin-induced agglutination.

Increased platelet reactivity to prostaglandin E1 in hypertension is linked with altered signal transduction.

Prostaglandin E1 has been shown to induce a greater accumulation of cAMP in platelets from spontaneously hypertensive (SHR) than in platelets from normotensive (Wistar-Kyoto, WKY) rats (Circ. Res. 1978;43:583-591. Thromb. Res. 1988;49:5-21). This study was conducted to determine the mechanisms of increased platelet reactivity to PGE1 in hypertension. The number of PGE1 binding sites/platelet (WKY 280 +/- 8, SHR 287 +/- 5) as well as the Kd for WKY (105 +/- 11 nmol/L) and SHR (120 +/- 14 nmol/L) were found to be similar in WKY and SHR rats. PGE1-induced GTPase activity was determined using WKY and SHR platelet membranes. The basal GTPase activity was similar in WKY and SHR platelets. Incubation of membranes with PGE1 (3 mumol/L) for 1 min increased GTPase activity by 46% in WKY and by 806% in SHR. Incubation of platelets with 1.0 mmol/L IBMX (3-isobutyl-1-methyl-xanthine) for 4 min resulted in a 327 +/- 57% and 320 +/- 11% increase in cAMP in WKY and SHR platelets, respectively. In other experiments, incubation of platelets with 3, 10 and 100 mumol/L forskolin, induced similar increases in cAMP levels in WKY (103 +/- 12%, 530 +/- 42%, 784 +/- 41%) and SHR (111 +/- 13%, 461 +/- 18%, 756 +/- 28%) platelets. These data lead us to suggest that the greater accumulation of cAMP induced by PGE1 in SHR than in WKY platelets is linked with altered PGE1-receptor mediated signal transduction at the G-protein level.

Protein prenylcysteine analog inhibits agonist-receptor-mediated signal transduction in human platelets.

Signal transduction components, including the Ras superfamily of low molecular weight GTP-binding proteins and the gamma subunits of heterotrimeric G proteins, are reversibly carboxyl methylated at C-terminal prenylcysteine residues. We have previously shown that the prenylcysteine analog N-acetyl-S-trans,trans-farnesyl-L-cysteine (AFC) inhibits carboxyl methylation of these proteins in human platelets. Here we show that concentrations of AFC that inhibit Ras carboxyl methylation (10-50 microM) also block responses to agonists such as ADP, collagen, arachidonic acid, U46619 (a stable analog of prostaglandin H2), thrombin, and guanosine 5'-[gamma-thio]triphosphate. AFC does not inhibit aggregation induced by effectors such as ionomycin, phorbol 12,13-dibutyrate, and bacterial phospholipase C that bypass G proteins to activate platelets at the level of cytosolic Ca2+ concentration and protein kinase C. These findings indicate that AFC inhibits agonist-receptor-mediated signal transduction in human platelets.

Normal thrombin binding leads to greater fibrinogen binding and increased platelet aggregation in spontaneously hypertensive rats.

This study was conducted to determine the mechanisms of increased platelet reactivity to thrombin in hypertension. Thrombin induced significantly greater platelet aggregation in spontaneously hypertensive (SHR) than in normotensive (Wistar Kyoto, WKY) rats. Fibrinogen and thrombin binding to platelets was determined using [125I]-fibrinogen and [125I]-thrombin respectively. Increased platelet aggregation in SHR correlated with thrombin-induced greater binding of fibrinogen to SHR than to WKY platelets. However, the number of thrombin receptors (binding sites/platelet) in WKY (19,500 +/- 3,000) and SHR (23,100 +/- 3,000) as well as thrombin dissociation constants were statistically similar in WKY (1.17 +/- 0.2 microM) and SHR (1.62 +/- 0.27 microM) platelets. Fura 2/AM, a fluorescent calcium indicator, loaded platelets were used to quantify the platelet ionized calcium ([Ca2+]i). The [Ca2+]i in unstimulated SHR and WKY platelets was essentially the same. In a calcium poor medium, thrombin-induced a 35% greater increase in [Ca2+]i in SHR than in WKY platelets. These data, taken together with our earlier observations that thrombin induces a significantly greater hydrolysis of phosphoinositide (Thromb. Res. 49, 5-21, 1988), lead us to suggest that thrombin-induced increased generation of inositol 1,4,5-trisphosphate and diacylglycerol induces greater fibrinogen binding and consequently increased aggregation in SHR than WKY platelets. The finding that the thrombin binding isotherms are similar in WKY and SHR platelets suggests that increased platelet sensitivity to thrombin in hypertension may be due to altered signal transduction and not due to changes in the number or affinity of thrombin receptors.

Interactions of La3+ with phosphatidylserine vesicles. Binding, phase transition, leakage and fusion.

The interaction of La3+ with phosphatidylserine vesicles is elucidated by binding studies, differential scanning calorimetry, X-ray diffraction, freeze fracture electron microscopy, and release of vesicle contents. La3+ effectively competes with Ca2+ for phosphatidylserine binding sites. The saturation level is close to a La/lipid ratio of 1:3. A concentration of 0.1 mM of La3+ is sufficient to induce fusion between sonicated vesicles.

Alternate reasons for atherogenesis in uremia.

The key to symptomatology in uremia is nitrogen retention leading to amidination and transmidination of a variety of substrates. The product of this activity is a series of guanidino acids which are methyl receptors converting S-adenosylmethionine to adenosine and homocysteine. Adenosine is a potent inhibitor of the enzyme ATPase and, in this way, contributes to the anemia, the bleeding diathesis and the CNS symptoms of uremia. Homocysteine is an inhibitor of pyridoxal phosphate-induced reactions and contributes to the angiitis and thromboembolism so unexpectedly encountered in chronic uremia.