Effect of interferon-alpha therapy on epitope-specific cytotoxic T lymphocyte responses in hepatitis C virus-infected individuals.

The majority of hepatitis C virus (HCV)-infected individuals fail to resolve the infection and become chronically infected despite the presence of HCV-specific CTL responses directed to different HCV-derived peptide antigens. Only a minority of individuals is able to clear the virus by mounting efficient CTL responses early after acute infection, but at present it is not clear whether viral clearance is associated with CTL responses of defined specificity. To elucidate those responses associated with improvement of the disease, we analyzed CTL responses to 16 different HLA-A2-presented, HCV-derived epitopes in 12 chronically infected patients, 14 chronically infected patients treated with interferon-alpha, and in one patient with acute symptomatic disease. We show here that the majority of chronically infected individuals present CTL responses directed to an NS4-derived peptide antigen (amino acids 1789-1797). Treated patients presented stronger HCV-specific CTL responses and therapy-induced changes in CTL target choice. In particular, 13 out of 14 individuals responded to an NS3-derived epitope (amino acids 1073-1081). By longitudinal analysis we show that five individuals responding to IFN-alpha therapy with decreases in alanine aminotransferase levels presented a strong CTL activity directed to the NS3-derived epitope. One patient that spontaneously resolved the infection presented a generally strong CTL activity specific for HCV-derived epitopes with a dominant response to the NS3-derived peptide antigen. This suggests that CTL responses directed to this NS3-derived antigen may be beneficial for the control of HCV infection. Improvement of these responses may represent a therapeutic intervention in chronic HCV infection.     

A comprehensive in vitro characterization of pancreatic ductal carcinoma cell line biological behavior and its correlation with the structural and genetic profile

There are a large number of stable pancreatic ductal carcinoma cell lines (PDCL) that are used by researchers worldwide. Detailed data about their differentiation status and genetic alterations are present in the literature, but a systematic correlation with cell biological behavior is often lacking. PDCL (n=12) were clustered by source of tumor cell (ascites, primary tumor, metastasis), and the data of functional cell biology were correlated with the reported structural and genetic profiles. Major histocompatibility complex expression, chemosensitivity and aneuploidia appeared to be related to the source of PDCL, and proliferative capacity appeared to be related to the grade of differentiation. No correlation between genetic/structural features of PDCL and biological behavior was found. All the cell lines appeared generally insensitive to in vitro treatment with 5-fluorouracil and showed variable degrees of susceptibility to gemcitabine, raltitrexed and oxaliplatin. All the PDCL showed resistance to Fas-mediated apoptosis but were significantly sensitive to the pro-apoptotic effect of inflammatory cytokines [interleukin (IL)-1, tumor necrosis factor (TNF) and interferon ]. PDCL were characterized for the secretion of several factors relevant to the tumor-immune cross talk. Vascular endothelial growth factor, CCL2, CCL5 and transforming growth factor were the factors most frequently released; less frequent was the secretion of CXCL8, CCL22, IL-6 and sporadically CXCL12, IL-10 and hepatocyte growth factor. The cytokines IL-1 and TNF were always undetectable. In conclusion, a clear correlation between structural/genetic features and function could not be detected, suggesting the weakness of a morphological classification for the in vitro studies of pancreatic cancer.     

Environmental enrichment prevents effects of dark-rearing in the rat visual cortex

Environmental enrichment potentiates neural plasticity, enhancing acquisition and consolidation of memory traces. In the sensory cortices, after cortical circuit maturation and sensory function acquisition are completed, neural plasticity declines and the critical period 'closes'. In the visual cortex, this process can be prevented by dark-rearing, and here we show that environmental enrichment can promote physiological maturation and consolidation of visual cortical connections in dark-reared rats, leading to critical period closure.     

Vasculogenic mimicry has no prognostic significance in pT3 and pT4 cutaneous melanoma

The concept of vasculogenic mimicry has been introduced to define periodic acid-Schiff (PAS)-positive channels and loops lined by tumor cells, instead of endothelium, able to contribute to microcirculation in uveal melanomas. Previous studies have shown that the PAS-positive patterns are associated with a poor prognosis in uveal melanoma. The aim of the current study was to investigate whether vasculogenic mimicry has a prognostic impact in pT3 and pT4 cutaneous melanoma. Fifteen patients with pT3 and pT4 cutaneous melanoma who did not experience progression after 10 years of follow-up and 30 matched controls who underwent progression were selected. Tumor sections were stained with PAS reaction, omitting the nuclear counterstaining. For immunohistochemistry, sections were stained with CD31, CD105 (endoglin), and laminin. Differences in the distribution of the PAS-positive patterns and a series of clinicopathological variables were evaluated by the Pearson chi(2) and Mann-Whitney U tests. We observed PAS-positive linear sheets, arcs, elliptical loops, and networks encircling roundish to oval aggregates of melanoma cells. The overall distribution of the PAS-positive patterns did not match with the blood microvessels' architecture as detected by immunohistochemical analysis. No statistically significant differences in the distribution of PAS-positive patterns were found between cases and controls. The presence of a parallel pattern correlated significantly with thickness (P = 0.04), whereas an inverse correlation was found with vessel area (P = 0.05). In conclusion, our results suggest that there is a mismatch between vasculogenic mimicry and tumor angiogenesis and do not support any prognostic role of vasculogenic mimicry in thick cutaneous melanoma.     

Reduced folate carrier polymorphism (80A-->G) and neural tube defects

Transport of folates in mammalian cells occurs by a carrier-mediated mechanism. The human folate carrier (RFC-1) gene has been isolated and characterized. Within this gene, a common polymorphism, 80A-->G, changing a histidine to an arginine in exon 2 (H27R), was recently identified. Defects in folate metabolism, such as defective carrier molecules, could be implicated in the etiology of neural tube defects (NTDs). In the present case-control study, we recruited 174 Italian probands with nonsyndromic NTD, 43 mothers, 53 fathers and 156 control individuals and evaluated the impact of RFC-1 variant on NTD risk. A statistically significant risk was calculated for the 80GG genotype of the NTD cases (OR=2.35; 95% CI 1.21-4.58) and mothers (OR=2.74; 95% CI 0.92-8.38). On the contrary, the heterozygous genotype of the mothers and both heterozygous and homozygous genotypes of the fathers did not seem to be significant NTD risk factors. Furthemore, according to the multifactorial inheritance of NTDs, we demonstrated that the combined genotypes for MTHFR 1298A-->C and RFC-1 80A-->G polymorphisms of cases resulted in greater NTD risk than heterozygosity or homozygosity for RFC-1 80A-->G variant alone. Conversely, our data provide no evidence for an association between NTD phenotype and combined MTHFR C677T/RFC-1 A80G genotypes. Moreover, here we describe the combinations of the two MTHFR polymorphic sites (677CT and 1298AC) with RFC-1 genotypes. We found that both patients and controls could have at most quadruple-mutation combinations. Interestingly, 27% (7/26) of the mothers and 18.75% (30/160) of the cases genotyped presented four mutant alleles in comparison with 8.5% (11/129) of the controls. Finally, the frequency of NTD cases and mothers carrying combined heterozygosity for the two MTHFR polymorphisms and RFC-1 80GG homozygosity (677CT/1298AC/80GG) (cases=11.3%; mothers 11.5%) was increased compared with controls (1.6%). Altogether, our findings support the hypothesis that RFC-1 A80G variant may contribute to NTD susceptibility in the Italian population.     

HER-2/neu and bcl-2 in ovarian carcinoma: clinicopathologic, immunohistochemical, and molecular study in patients with shorter and longer survival.

The bcl-2 protein is a membrane protein involved in prolonging cell survival by inhibiting apoptosis. The HER-2 oncogene, which is located on chromosome 17 and encodes for a tyrosine-kinase growth factor receptor, is amplified and HER-2/neu is overexpressed in 25% to 30% of breast carcinomas. The authors analyzed the bcl-2 expression and the bcl-2 gene and HER-2/neu overexpression and amplification in FIGO stage IIIC, serous, G3, ovarian carcinomas obtained from living patients who had no evident disease 5 years after primary treatment compared with ovarian carcinomas obtained from patients, matched for stage, grade of differentiation, and treatment, who had died of progression of disease no later than 2 years after primary treatment. bcl-2 overexpression was statistically correlated with progression of disease during first-line chemotherapy (P=0.021). The HER-2/neu status was found not to correlate with progression of disease during first-line chemotherapy. Both bcl-2 and HER-2/neu expression were not statistically associated with the clinical outcome of ovarian cancer patients. Gene amplification of the HER-2/neu chromosome 17 was found in all the HER-2/neu, 3+ score, positive-staining ovarian carcinomas. None of the analyzed samples revealed a translocation t(14;18)(q32;q21) in the bcl-2 gene. The knowledge of additional prognostic or even predictive factors, such as bcl-2 expression, in patients with advanced ovarian carcinoma before the primary chemotherapeutic treatment may help in the management of patients who require a more tailored treatment. In addition, the gene amplification of the HER-2/neu suggests that HER-2 is a potential target for treatment in ovarian cancer.     

Correlation of epidermal growth factor receptor expression with tumor microdensity vessels and with vascular endothelial growth factor expression in ovarian carcinoma

We analyzed in advanced ovarian serous G3 carcinoma the correlation between epidermal growth factor receptor (EGFR) overexpression and tumor angiogenesis and their relation with clinical outcome. Microvessel density (MVD) and vascular endothelial growth factor (VEGF) were statistically correlated with disease-free interval and death from disease both in univariate and multivariate analyses while EGFR expression was not correlated with clinical outcome. MVD was significantly associated with progression of disease during chemotherapy while VEGF and EGFR expression were not correlated with responsiveness to chemotherapy (Fisher's exact test). VEGF expression was correlated with MVD (Fisher's exact test). EGFR showed a trend to correlation with MVD. Further studies focusing on the use of angiogenesis inhibitors in addition to EGFR inhibitors on ovarian carcinoma cells may produce therapeutic strategies in the selection of tailored therapies in ovarian cancer patients.     

Effects of soy isoflavones on endothelial function in healthy postmenopausal women

OBJECTIVE: To evaluate the effects of soy isoflavone administration on endothelial function in healthy postmenopausal women. DESIGN: Sixty naturally postmenopausal women were randomly assigned to receive isoflavone or placebo tablets for 6 months. Endothelium-dependent vasodilatation was measured by brachial reactivity technique along with levels of plasma soluble intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), E-selectin, P-selectin and soluble thrombomodulin, von Willebrand factor, and tissue plasminogen activator. Differences between endothelium-dependent and endothelium-independent vasodilatation were assessed by evaluating brachial reactivity parameters after reactive hyperemia and after sublingual administration of nitroglycerin; furthermore, in the active group, the effect of isoflavones was also evaluated during the intra-arterial infusion of N-monomethyl-L-arginine. Serum levels of lipids [high-density lipoprotein and low-density lipoprotein cholesterol, triglycerides and lipoprotein(a)] and hemostatic factors (prothrombin, fibrinogen, plasminogen activator inhibitor-1, and fibrin D-dimer) were also measured. To confirm the absorption of isoflavones, their blood concentrations were determined. RESULTS: Isoflavone treatment versus placebo was associated with a significant improvement in endothelium-dependent vasodilatation but had no impact on endothelial-independent arterial diameter and flow. Intra-arterial infusion of N-monomethyl-L-arginine inhibited the significant effect of isoflavones on endothelium-mediated vasodilatation. Furthermore, isoflavone group experienced statistically significant reductions in plasma concentrations of ICAM-1, VCAM-1, and E-selectin. Levels of soluble thrombomodulin, von Willebrand factor, tissue plasminogen activator, lipids, and hemostatic factors did not change significantly throughout the study in both groups. CONCLUSIONS: Our findings suggest a positive influence of soy isoflavones on endothelial function in healthy postmenopausal women as evidenced by an improvement in endothelium-dependent vasodilatation and a reduction in plasma adhesion molecule levels.     

Alteration of acylphosphatase levels in familial Alzheimer''s disease fibroblasts with presenilin gene mutations

Acylphosphatase (ATPase), an enzyme that modulates the activity of Ca²⁺-ATPase by hydrolysing its phosphorylated moiety, has been found to be significantly higher in cultured skin fibroblasts from donors affected by early onset familial Alzheimer's disease (EOFAD) with PS-1 and PS-2 gene mutations. Of the two known isoenzymes of acylphosphatase, only the erythrocyte one accounts for the total increase in activity. No relevant alteration was observed in phosphotyrosine phosphatase activity (PTPase), in Ca²⁺-ATPase and Na⁺,K⁺-ATPase activities of the same cells as compared to age-matched controls. This finding could suggest a possible explanation for the calcium-dependent biochemical alterations previously described in Alzheimer's disease fibroblasts.