Human immunodeficiency virus (HIV) distribution in HIV encephalitis: study of 19 cases with combined use of in situ hybridization and immunocytochemistry.

Brains of 19 AIDS patients with HIV encephalitis were examined by immunohistochemistry and in situ hybridization using antisense HIV DNA and RNA probes. Double immunohistochemical labeling, using antibodies against viral and cell-type specific antigens, was utilized to study lesions in some brains. Other combined studies included use of in situ hybridization and immunohistochemical labeling of the same section, using antibodies against either viral or cell-type specific antigens. Hybridization signals were abundant and were concentrated mainly in the white matter. Heavy labeling was found in the subcortical white matter, the corpus callosum, the internal capsule, and white matter regions of the brainstem and cerebellum. Deeper cortical layers often contained cells with hybridized probe when the subcortical white matter was intensely labeled. HIV nucleic acid sequences were found almost exclusively in macrophages. Counts showed that 16-25% of macrophages contained viral antigens and exhibited hybridized HIV probe. Almost all of these macrophages contained proviral DNA, viral RNA and viral proteins; i.e. they were actively replicating HIV. We also examined brains from three AIDS cases without clinical or pathological evidence of HIV encephalitis; no HIV sequences or immunoreactive proteins were detected.

Search for HIV proviral DNA and amplified sequences in the muscle biopsies of patients with HIV polymyositis.

We searched for the presence of human immunodeficiency virus (HIV) in fresh-frozen muscle biopsy specimens from 10 patients with HIV-associated polymyositis (HIV-PM) using (a) 35S-labeled HIV-RNA transcript of the virus and in situ hybridization, and (b) polymerase chain reaction and slot-blot hybridization utilizing primers amplifying sequences from the gag and pol genes of the HIV genome. With in situ hybridization, positive signals were detected in sparse lymphoid cells surrounding the muscle fibers, but not within the muscle fibers, in up to two consecutive sections in 6 of the 10 specimens. By the polymerase chain reaction, amplified HIV-specific sequences were noted in 2 specimens, but in only 2 of 8 consecutive sections, implying infection of lymphoid cells rather than muscle fibers. Muscle cultures from six specimens failed to show integrated HIV sequences within the myotubes. We conclude that HIV sequences or transcriptional products are not present within the muscle fibers or the cultured myotubes of patients with HIV-PM. This indicates that: (a) viral replication does not take place within the muscle; (b) integration of HIV proviral genome does not occur within the myonuclei or satellite cells; and (c) HIV-PM does not seem to be due to a persistent infection of the muscle fiber by the virus.

Replication of HIV-1 and HIV-2 in human bone marrow cultures.

The bone marrow is a target organ for the human immunodeficiency virus (HIV), but the mechanisms by which suppression of hematopoiesis occurs during the course of HIV infection are not well understood. To study this issue, the effect of several different HIV-1 isolates (monotrophic and lymphotrophic) and one HIV-2 isolate on in vitro colony formation by BFU-E and CFU-GM from normal human marrow were examined. The monotrophic strain AD-87 (M) failed to replicate in marrow cultures as documented by RT, and colony formation by BFU-E and CFU-GM was unaffected by this virus. A derivative of this isolate AD-87 (M-P), which was replicated in peripheral blood lymphocytes (PBL), however, replicated well and markedly inhibited colony formation by BFU-E and CFU-GM. Two additional PBL isolates replicated less efficiently; neither inhibited CFU-GM but one consistently inhibited BFU-E colony formation. Inhibition of colony formation by the HIV-1 isolates was a late event, presumably a secondary lysis of cells, since up to 7 days after inoculation colony numbers were normal but diminished markedly by 10 days, and since only up to 10% of the cells of the monocyte lineage contained detectable virus by in situ, EM, and IFA studies. In contrast, the HIV-2 isolate was so lytic that by 4 days after inoculation the majority of the marrow cells were destroyed.

Ultraviolet radiation increases HIV-long terminal repeat-directed expression in transgenic mice.

Previously described FVB/N mice harboring a human immunodeficiency virus (HIV) long terminal repeat (LTR)/chloramphenicol acetyl transferase (CAT) transgene were treated with varying amounts of 254 nm UV-C radiation or 312 nm UV-B radiation. At optimal exposure periods, a 20-fold increase in HIV-LTR-directed expression was observed in ear specimens collected 24 h following UV-C exposure; a fourfold increase in expression was induced by UV-B exposure. Investigation of the kinetics of UV-C induction in vivo revealed that LTR-directed gene expression began to increase 2 hours after exposure and reached a maximum on Day 3 following exposure (greater than 30-fold induction). In experiments examining the kinetics of UV-B activation, the maximum level of CAT activity in the ears of irradiated transgenic animals was fivefold above levels in unirradiated transgenic controls (Day 5). Furthermore, CAT activity was not induced in fur-bearing skin following UV exposure; however, a fourfold increase in HIV-LTR-directed expression could be elicited when hair was removed by shaving prior to UV-B treatment.

Development of disease and virus recovery in transgenic mice containing HIV proviral DNA.

Transgenic mice containing intact copies of the human immunodeficiency virus (HIV) proviral DNA were constructed. Founder animals were not viremic for HIV and remained healthy during a 9-month observation period. After being mated with nontransgenic animals, one founder mouse (No. 13) gave rise to F1 progeny that developed a disease syndrome characterized by marked epidermal hyperplasia, lymphadenopathy, splenomegaly, pulmonary lymphoid infiltrates, growth retardation, and death by day 25 of life. Infectious HIV, indistinguishable from parental virus by immunoblot analysis, was recovered from the spleen, lymph nodes, and skin of five of five affected animals.