No abstract

To investigate the effect of HIV‐specific CD8+ T cells on viral plasma load and disease progression, we enumerated HLA‐A2‐, B8‐ and B57‐restricted CD8+ T cells directed against several HIV epitopes in a total of 54 patients by the use of tetrameric HLA‐peptide complexes. In patients with high CD4+ T cell numbers, HIV‐specific tetramer+ cells inversely correlated with viral load. Patients with CD4+ T cell numbers below 400/μ l blood, however, carried high viral load despite frequently having high tetramer+ T cell numbers. This lack of correlation between viral load and tetramer+ cells did not result from viral escape variants, as in only 4 of 13 patients, low frequencies of viruses with mutated epitopes were observed. In 15 patients we measured CD8+ T cell antigen responsiveness to HIV peptide stimulation in vitro. FACS analyses showed differential IFN‐γ production of the tetramer+ cells, and this proportion of IFN‐γ‐producing tetramer+ cells correlated with AIDS‐free survival and with T cell maturation to the CD27– effector stage. These data show that most HIV‐infected patients have sustained HIV‐specific T cell expansions but many of these cells seem not to be functional, leaving the patient with high numbers of non‐functional virus‐specific CD8+ T cells in the face of high viral burden.

Crigler-Najjar (CN) patients have no bilirubin UDP glucuronosyltransferase (UGT1A1) activity and suffer brain damage because of bilirubin toxicity. Vectors based on adeno-associated virus (AAV) serotype 2 transduce liver cells with relatively low efficiency. Recently, AAV serotypes 1, 6, and 8 have been shown to be more efficient for liver cell transduction. We compared AAV serotypes 1, 2, 6, and 8 for correction of UGT1A1 deficiency in the Gunn rat model of CN disease. Adult Gunn rats were injected with CMV-UGT1A1 AAV vectors. Serum bilirubin was decreased over the first year by 64% for AAV1, 16% for AAV2, 25% for AAV6, and 35% for AAV8. Antibodies to UGT1A1 were detected after injection of all AAV serotypes. An AAV1 UGT1A1 vector with the liver-specific albumin promoter corrected serum bilirubin levels but did not induce UGT1A1 antibodies. Two years after injection of AAV vectors all animals had large lipid deposits in the liver. These lipid deposits were not seen in age-matched control animals. AAV1 vectors are promising candidates for CN gene therapy because they can mediate a reduction in serum bilirubin levels in Gunn rats that would be therapeutic in humans.

The ATP binding cassette transporters ABCG5 and ABCG8 are indispensable for hepatobiliary cholesterol transport. In this study, we investigated the specificity of the heterodimer for cholesterol acceptors. Dog gallbladder epithelial cells were mono-or double-transfected with lentiviral mouse Abcg5 and Abcg8 vectors. Double-transfected cells showed increased efflux to different bile salt (BS) species, while mono-transfected cells did not show enhanced efflux. The efflux was initiated at micellar concentrations and addition of phosphatidylcholine increased efflux. Cholesterol secretion was highly BS dependent, whereas other cholesterol acceptors such as ApoAI, HDL or methyl-b-cyclodextrin did not elicit Abcg5/g8 dependent cholesterol secretion.

HLA‐B57 has been shown to be associated with long‐term asymptomatic HIV‐1 infection. To investigate the biological mechanism by which the HLA‐B57 allele could protect from HIV‐1 disease, we studied both the number of CD8+ T cells as well as CD8+ T cell responsiveness directed to different HIV‐1 Gag peptides presented by HLA‐A2, ‐B8 or ‐B57. T cells specific for the HLA‐B57 peptide KAFSPEVIPMF responded more readily and to a higher extend to antigenic stimulation in vitro than T cells specific for the HLA‐A2 peptide SLYNTVATL or the HLA‐B8 peptide EIYKRWII. This phenomenon was reproducible with T cells from individuals expressing HLA‐B57 in combination with one or both of the other alleles and was persistent during long‐term follow‐up. Lower reactivity of A2‐ and B8‐restricted T cells was not explained by mutations in the B8‐ or A2‐restricted Gag‐peptides. Moreover, no correlation between peptide mutation frequency and IFN‐γ production by the corresponding Gag‐specific T cells was observed. In conclusion, functional differences were observed between T cells specific for HIV epitopes derived from the same protein presented by different HLA molecules. B57‐restricted KAFSPEVIPMF‐specific CD8+ T cells have relatively high responsiveness, which could contribute to the protective effect of HLA‐B57 in HIV infection.

FABACs (fatty acid-bile acid conjugates) are synthetic molecules that are designed to treat a range of lipid disorders. The compounds prevent cholesterol gallstone formation and diet-induced fatty liver, and increase reverse cholesterol transport in rodents. The aim of the present study was to investigate the effect of FABACs on cholesterol efflux in human cells. Aramchol (3beta-arachidylamido-7alpha,12alpha,5beta-cholan-24-oic acid) increased cholesterol efflux from human skin fibroblasts in a dose-dependent manner in the absence of known efflux mediators such as apoA-I (apolipoprotein A-I), but had little effect on phospholipid efflux. An LXR (liver X receptor) agonist strongly increased Aramchol-induced cholesterol efflux; however, in ABCA1 (ATP-binding cassette transporter A1)-deficient cells from Tangier disease patients, the Aramchol effect was absent, indicating that activity of ABCA1 was required. Aramchol did not affect ABCA1 expression, but plasma membrane levels of the transporter increased 2-fold. Aramchol is the first small molecule that induces ABCA1-dependent cholesterol efflux without affecting transcriptional control. These findings may explain the beneficial effect of the compound on atherosclerosis.