Longitudinal study in HIV/HCV-coinfected HAART-naive patients and role of HCV genotype.

To evaluate the impact of highly active antiretroviral therapy (HAART) on the course of hepatitis C (HCV) infection, we studied the biological and virological characteristics of 23 HCV/HIV-coinfected HAART-naive patients. The HCV genotype, HCV and HIV viral loads, serum alanine aminotransferase, CD4+ and CD8+ cell/mm3 were determined at baseline, 1 month, 6 months and 12 months after initiation of HAART. Results were analyzed both in terms of total population and of HCV genotype. The study of the total population suggests that this therapy did not determine a significant alteration of HCV viremia and levels of ALT, while a significant decrease in HIV viremia (-1.7log10 at one year from the start of HAART) and increase in CD4+ counts was observed (P < 0.005). The biological and virological parameters of HCV/HIV coinfection differed according to the HCV genotype. In particular, only genotype 4 showed a significant inverse correlation between HCV and HIV viral loads.

Vertical transmission of the hepatitis C virus to infants of anti-human immunodeficiency virus-negative mothers: molecular evolution of hypervariable region 1 in prenatal and perinatal or postnatal infections.

In a prospective study of 33 infants born to hepatitis C virus (HCV)-positive human immunodeficiency virus-negative mothers the vertical transmission of HCV occurred in 6.8%. The evolution of HCV infection in two babies was studied from birth up to 5 or 6 years of age, and the sequencing of the hypervariable region (HVR) of the putative envelope-encoding E2 region of the HCV genome was performed. The HVR1 sequence variability and the different serological profiles during follow-up could reflect the differences in HCV transmission routes, HCV genotypes, and clinical evolution of infection.

Differential expression of antiapoptotic genes in human endometrial carcinoma: bcl-XL succeeds bcl-2 function in neoplastic cells.

OBJECTIVES: Previous histochemical observations have suggested a possible involvement of the bcl-2 family genes in the acquisition of neoplastic phenotype of the endometrium. Since knowledge of the type and function of genes controlling the transformed cell may result in new diagnostic, prognostic, and therapeutic approaches, we have investigated at the molecular level the biological role of bcl-2 family genes in endometrial neoplastic cells. METHODS: To investigate the relationship between the sensitivity to apoptosis and the expression of the bcl-2 family genes, we set up a model system consisting of four human endometrial carcinoma cell lines. This system constitutes an array of two cell pairs presenting, respectively, endometrioid and adenosquamous phenotypes. G2 and G3 gradings are represented within each pair; in addition, each set contains one cell line that is apoptosis-sensitive and one that is resistant. Transfection of bcl-2 and bcl-XL into apoptosis-sensitive cells was used to monitor the biological function of protective genes. RESULTS: A differential pattern of expression of bcl-2 family genes was observed in apoptosis-sensitive versus resistant cells, independent from the histological subtype. Resistant lines exhibited high amounts of Bcl-XL and low amounts of Bcl-2. Bax expression clearly correlates with cellular susceptibility to apoptosis. Transfection of bcl-XL resulted in a dose-dependent enhancement in resistance toward apoptosis. In contrast, the main effect of bcl-2 constitutive overexpression was to drastically abate the proliferative potential of transfected cells. CONCLUSIONS: These data demonstrate, at the molecular level, that bcl-XL is selected as an apoptosis-protective gene in place of bcl-2 while bax retains its dominant proapototic role.

Decreased phosphorylation of mutant insulin receptor by protein kinase C and protein kinase A.

We have recently reported that the Arg1152-->Gln insulin receptor mutation (QK single mutant) alters a conserved motif (RK motif) immediately next to the key tyrosine phosphorylation sites (Tyr1146, Tyr1150, Tyr1151) of the receptor and constitutively activates its kinase and metabolic signaling. To investigate further the function of the RK motif, we have expressed two additional mutant insulin receptors: a single mutant, in which the second basic residue in the RK motif (Lys1153) was substituted (RA mutant); and a double mutant, in which both the Arg and the Lys residues were replaced with noncharged amino acids (QA mutant). As compared with the transfected wild-type receptors (WT), both the single and the double mutant receptors were normally synthetized and transported to the plasma membrane and bound insulin normally. Whereas the double mutant receptor exhibited preserved insulin-dependent autophosphorylation, kinase activity, and 2-deoxyglucose uptake, all of these functions were grossly impaired in the two single mutant receptors. Two-dimensional analysis of tryptic phosphopeptides from receptor beta-subunits revealed that decreased autophosphorylation of the single mutant receptors mainly involved regulatory Tyr1150,1151 and carboxyl-terminal Tyr1316,1322. At variance with the insulin-stimulated, insulin-independent tyrosine kinase activity toward poly(Glu-Tyr) 4:1 was increased 3-fold in both the double and the single mutants. All mutant receptors induced a 2-fold increase in basal 2-deoxyglucose uptake in NIH-3T3 cells. Treatment of WT transfected cells with 12-O-tetradecanoyl-phorbol-13-acetate or 8-bromo-cAMP increased insulin receptor phosphorylation by 3-fold. No phosphorylation was observed in cells expressing the two single or the double mutant receptor. Consistently, purified preparations of PKC and PKA phosphorylated the WT but not the mutant receptors in vitro. A 17-amino acid synthetic peptide encoding the receptor sequence surrounding the RK motif inhibited phosphorylation of WT insulin receptors by both protein kinases A and C. A mutant peptide in which the RK sequence was replaced by QK (to mimic the mutation in the QK receptor) exhibited no inhibitory effect. Thus, the RK insulin receptor motif is required for insulin receptor phosphorylation by protein kinases C and A and may modulate insulin-independent receptor activity. The RK motif may also have an important structural role in allowing normal insulin regulation of the kinase.