PDGFRalpha, PDGFRbeta and KIT expression/activation in conventional chondrosarcoma.

Chondrosarcomas represent 20% of all primary bone sarcomas, and many studies have attempted to unravel molecular targets for future development of new therapies. The aim of this study was to investigate the expression/activation of PDGFRalpha, PDGFRbeta and KIT receptor tyrosine kinases (RTKs) as potential therapeutic targets in conventional central primary chondrosarcomas (CCS). The expression of PDGFRalpha, PDGFRbeta and KIT RTKs was detected in 16 CCSs using immunohistochemistry (IHC), and their level of expression and activation status were analysed by immunoprecipitation and western blot experiments. PDGFRalpha, PDGFRbeta and KIT cDNAs were screened to verify the presence of activating mutations and the presence of the cognate ligands was analysed by means of RT-PCR. RTK gene amplification was further studied by means of fluorescence in situ hybridization (FISH) analysis. The immunophenotyping and biochemical analyses showed that the CCSs co-expressed PDGFRalpha and PDGFRbeta, with the latter showing definitively greater protein expression and phosphorylation levels. PDGFRbeta was expressed but not activated in control healthy joint cartilage, in line with no PDGFB detection. Conversely, the KIT gene product did not seem to play a relevant role. These findings, in the absence of activating mutations or an abnormal genomic profile and the presence of PDGFA and PDGFB expression, are consistent with an autocrine/paracrine loop activation of the corresponding receptors. The CCS gene profile described here offers a rationale for the use of RTK inhibitors alone or in combination with chemotherapy, and supports further investigation of RTKs and their downstream signals.

Pro-oxidant activity and methionine metabolism in chronic alcohol abusers: relationship to alcohol withdrawal and folate administration.

AIM: The aim of this study was to investigate whether alcohol withdrawal and folate administration could play a role on redox balance and metionine metabolism in heavy drinkers. METHODS: The derivatives of reactive oxygen metabolites (d-ROMs), homocysteine, total thiols, vitamin B12 and folate were evaluated in a selected group of 40 consecutive chronic alcohol abusers by comparison with 44 healthy moderate drinkers, as controls. RESULTS: Before alcohol withdrawal, d-ROMs were significantly higher (p<0.0001) in heavy drinkers than in controls: 368.5 (254.8-718.6) U.CARR vs 245 (200.7-360) U.CARR, respectively, median with range. Plasma homocysteine were significantly higher in alcoholics than in moderate drinkers (p<0.0001): 18 (9.5-82.2) micromol/L vs 9.1 (4.9-19.6) micromol/L, respectively. Heavy drinkers also exhibited higher serum thiols than moderate drinkers (p<0.003): 605.8 (448.2-717.7) micromol/L vs 554.8 (508.3-658.4) micromol/L, respectively. The patients showed lower plasma folate than controls (p<0.0001): 4.1 (1.9-9.7) ng/mL vs 8.8 (5.0-8.4) ng/mL, respectively, but similar vitamin B12 levels: 487 (299-786) pg/mL 621 (243-894) pg/mL. A negative correlation between homocysteine and folate was observed before withdrawal in alcoholics (r=-0.4546, p<0.038). Both serum thiols (549.7 micromol/L, range 402.4-616.6 micromol/L) and homocysteinemia (6.6 micromol/L, range 2.9-18.5 micromol/L) were significantly decreased (p<0.0001 and p<0.022, respectively) after a week of alcohol withdrawal and folate administration. CONCLUSION: Our findings show that both enhanced pro-oxidant activity and a derangement of methionine metabolism can be observed in heavy drinkers before alcohol withdrawal and folate administration. Furthermore, folate seems to be a strong determinant of both plasma homocysteine and thiol concentrations.

Increased erythrocyte glutathione peroxidase activity and serum tumor necrosis factor-alpha in HIV-infected patients: relationship to on-going prothrombotic state.

A condition of oxidative stress, due to perturbation of oxidant/antioxidant balance, has been suggested to play a role not only in the pathogenesis of human immunodeficiency virus (HIV) infection, but also in the promotion of a thrombophilic condition. Because various hemostatic dysfunctions usually considered as risk factors for thrombotic events were reported in HIV infection, this study was undertaken to investigate whether the oxidative phenomenon could promote a prothrombotic state in such condition. Erythrocyte glutathione peroxidase (GSH-Px), the major free-radical scavenger enzyme, and serum tumor necrosis factor-alpha (TNF-alpha) were evaluated in 33 consecutive HIV-infected out-patients and 35 matched HIV-negative healthy controls at a distance of any acute episode. Thrombin generation was explored by measuring the plasma levels of prothrombin fragment 1 + 2 (F1 + 2), whereas fibrin degradation products (D-dimer) and plasminogen activator inhibitor (PAI-1) activity were evaluated as indices of plasmin activity and fibrinolytic derangement. The anticoagulant pathway was investigated by measuring the plasma levels of antithrombin and protein C. Erythrocyte GSH-Px activity and serum TNF-alpha were significantly higher in HIV-infected patients when compared to controls. F1 + 2, D-dimer, and PAI-1 activity were increased in HIV-infected patients by comparison with controls. Normal antithrombin, but decreased protein C, was instead detected in HIV-infected patients. In the latter patients, serum TNF-alpha negatively correlated with both erythrocyte GSH-Px activity and plasma D-dimer. On the other hand, a positive correlation was shown between F1 + 2 and D-dimer and between D-dimer and GSH-Px activity. Furthermore, a trend toward increasing levels of GSH-Px with increasing PAI-1 activity was reported. These findings suggest a relationship between erythrocyte oxidative stress and the hypercoagulable condition during HIV infection.