No abstract

One of the major drawbacks limiting the use of synthetic peptide vaccines in genetically distinct populations is the fact that different epitopes are recognized by T cells from individuals displaying distinct major histocompatibility complex molecules. Immunization of mice with peptide (181-195) from the immunodominant 43 kDa glycoprotein of Paracoccidioides brasiliensis (gp43), the causative agent of Paracoccidioidomycosis (PCM), conferred protection against infectious challenge by the fungus. To identify immunodominant and potentially protective human T-cell epitopes in gp43, we used the TEPITOPE algorithm to select peptide sequences that would most likely bind multiple HLA-DR molecules and tested their recognition by T cells from sensitized individuals. The 5 most promiscuous peptides were selected from the gp43 sequence and the actual promiscuity of HLA binding was assessed by direct binding assays to 9 prevalent HLA-DR molecules. Synthetic peptides were tested in proliferation assays with peripheral blood mononuclear cells (PBMC) from PCM patients after chemotherapy and healthy controls. PBMC from 14 of 19 patients recognized at least one of the promiscuous peptides, whereas none of the healthy controls recognized the gp43 promiscuous peptides. Peptide gp43(180-194) was recognized by 53% of patients, whereas the other promiscuous gp43 peptides were recognized by 32% to 47% of patients. The frequency of peptide binding and peptide recognition correlated with the promiscuity of HLA-DR binding, as determined by TEPITOPE analysis. In silico prediction of promiscuous epitopes led to the identification of naturally immunodominant epitopes recognized by PBMC from a significant proportion of a genetically heterogeneous patient population exposed to P. brasiliensis. The combination of several such epitopes may increase the frequency of positive responses and allow the immunization of genetically distinct populations.

Background:The mechanisms regulating internalization of plasma membrane cholesterol in mammalian cells are not well understood. Results: A cell line haploinsufficient for U60 snoRNA expression exhibits impaired plasma membrane to ER cholesterol trafficking and increased de novo cholesterol synthesis. Conclusion: U60 snoRNA expression regulates cholesterol homeostasis by affecting internalization of plasma membrane cholesterol. Significance: This is the first study to implicate a snoRNA in regulation of cholesterol homeostasis.

The hypothesis of the role of iron overload associated with HFE gene mutations in the pathogenesis of nonalcoholic steatohepatitis (NASH) has been raised in recent years. In the present study, biochemical and histopathological evidence of iron overload and HFE mutations was investigated in NASH patients. Thirty-two NASH patients, 19 females (59%), average 49.2 years, 72% Caucasians, 12% Mulattoes and 12% Asians, were submitted to serum aminotransferase and iron profile determinations. Liver biopsies were analyzed for necroinflammatory activity, architectural damage and iron deposition. In 31 of the patients, C282Y and H63D mutations were tested by PCR-RFLP. Alanine aminotransferase levels were increased in 30 patients, 2.42 ± 1.12 times the upper normal limit on average. Serum iron concentration, transferrin saturation and ferritin averages were 99.4 ± 31.3 g/dl, 33.1 ± 12.7% and 219.8 ± 163.8 µg/dl, respectively, corresponding to normal values in 93.5, 68.7 and 78.1% of the patients. Hepatic siderosis was observed in three patients and was not associated with architectural damage (P = 0.53) or with necroinflammatory activity (P = 0.27). The allelic frequencies (N = 31) found were 1.6 and 14.1% for C282Y and H63D, respectively, which were compatible with those described for the local population. In conclusion, no evidence of an association of hepatic iron overload and HFE mutations with NASH was found. Brazilian NASH patients comprise a heterogeneous group with many associated conditions such as hyperinsulinism, environmental hepatotoxin exposure and drugs, but not hepatic iron overload, and their disease susceptibility could be related to genetic and environmental features other than HFE mutations.