Restriction of HIV-1 replication in intestinal cells is genetically controlled by the gag-pol region of the HIV-1 genome.

The human colon epithelial line HT29 represents a semipermisive cellular system for human immunodeficiency virus type 1 (HIV-1). It could be productively infected with HIV-1 NDK, a Zairian virus isolate highly cytopathic for CD4 positive lymphocytes, whereas infection with the prototype virus HIV-1 LAV was nonproductive. Recombinant viruses derived from HIV-1 LAV and HIV-1 NDK were used to determine the genetic control, step of virus/cell cycle, and molecular mechanism responsible for productive versus nonproductive infection of intestinal cells. Both parental viruses and all recombinants retrotranscribed their genomes with a similar kinetics and were able to complete HIV-1 DNA synthesis, HIV-1 LAV provirus present in preintegration complexes could be rescued by cocultivation with T-lymphocytes. However, it was aborted during prolonged cultivation of HT29 cells. Our results suggest that (i) gag/pol region of HIV-1 genome (fragment BssHII255-EcoRI4183) genetically controlled productive infection of intestinal cells and that (ii) the difference between productive and abortive infection occurred before synthesis of HIV-1 mRNA, at the integration level.

Characterization of HIV1-PAR, a macrophage-tropic strain: cell tropism, virus/cell entry and nucleotide sequence of the envelope glycoprotein.

The HIV1-PAR strain, isolated from the cerebrospinal fluid of an HIV1-seropositive man suffering from encephalopathy, replicated well in cord blood lymphocytes, poorly in peripheral blood mononuclear cells, and to different levels in blood-derived macrophage (BDM) cultures prepared from different blood donors. In marked contrast to its replication in primocultures, it did not grow in CEM and U937 cell lines. HIV1-PAR production in BDM was inhibited by more than 90% after treatment with OKT4A or 13B8.2 monoclonal antibodies (mAb) binding to adjacent epitopes of the D1 domain of the CD4 molecules. A lower but significant inhibitory effect was observed after BDM treatment with BL4 and OKT4 mAb, directed to the D2 and D3 domain of the CD4 molecule, respectively. The entire HIV1-PAR envelope glycoprotein gene was amplified by polymerase chain reaction and sequenced. The deduced amino acid sequence of HIV1-PAR gp160 revealed the presence of 847 amino acids and 86% homology with the HIV1 LAV virus prototype. An alignment of the amino acid sequence of the envelope glycoprotein of HIV1-PAR and HIV1-LAV showed that the differences were mostly clustered within the five variable regions. Five CD4-binding domains, the gp120/gp41 cleavage site, the putative gp41 fusion domain and 21 out of the 22 cysteine residues were conserved in both isolates. The results further confirm the macrophage-tropic character of the HIV1-PAR virus.

Inhibition of HIV by an anti-HIV protease synthetic peptide blocks an early step of viral replication.

The processing of the human immunodeficiency virus (HIV) gag and gag-pol precursor proteins by the virus-encoded protease is an essential step in maturation of infectious virus particles. Like most retroviral proteases, the HIV protease belongs to the aspartyl-protease family and can be inhibited by specific inhibitors. Twenty-four synthetic peptides known to be inhibitors of human renin were tested for inhibition of HIV replication in tissue cultures. One of them, a synthetic peptide analogue, SR41476, which has been shown to be a specific inhibitor of purified recombinant HIV1 protease in vitro, totally blocked infection with different isolates including the HIV1 LAV prototype, the highly cytopathic Zairian isolate HIV1 NDK, and HIV2 ROD, both in primary blood lymphocytes (PBL) and in the lymphoid cell lines MT4 and CEM, for at least 3 weeks. It also significantly reduced virus replication in chronically infected CEM cells, without any effect on cell proliferation. Radioimmunoprecipitation assay revealed that the inhibitor blocked processing of polyprotein precursors p55 gag and p40 gag into a mature form of gag proteins, p25 and p18. Synthetic peptide analogue SR 41476, when added before infection, efficiently inhibited formation of HIV DNA provirus and successfully suppressed synthesis of HIV-specific proteins. These results imply that the HIV protease inhibitor not only inhibited virus maturation in the late phase of the HIV replication cycle, but also interfered in the early phase, before the provirus was formed. This mechanism of antiviral activity provides new possibilities and strategies for AIDS chemotherapy.

Effects of the antiglucocorticoid RU 486 on the initiation of ultrastructural type-II cell differentiation in fetal rat lung.

The possible early role of endogenous glucocorticosteroids on the further ultra-structural development of the fetal rat lung epithelium was investigated in vitro using a potent antiglucocorticoid drug, RU 486. Lung primordia were explanted on day 13 of gestation and cultured for 4-6 days in the presence of fetal calf serum, with or without RU 486 in excess. The results obtained show that osmiophilic lamellar bodies specific for morphologically differentiated type-II cells did appear in a number of epithelial cells, even though the glucocorticosteroids possibly present in the culture medium, or transferred at explantation were antagonized by RU 486. The number of lamellar bodies stored in these pneumocytes was not different from that in controls. In contrast, their total surface area per cell profile was transiently decreased. Taken together the reported data strongly suggest that endogenous glucocorticosteroids are not necessary for the initiation of type-II cell differentiation.