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Biomedical subjects

J A Nelson

Publications and source records attributed to J A Nelson.

At least 73 records · Page 4Linked to original sources
PubMed1998

Exclusive and persistent use of the entry coreceptor CXCR4 by human immunodeficiency virus type 1 from a subject homozygous for CCR5 delta32.

Individuals who are homozygous for the 32-bp deletion in the gene coding for the chemokine receptor and major human immunodeficiency virus type 1 (HIV-1) coreceptor CCR5 (CCR5 -/-) lack functional cell surface CCR5 molecules and are relatively resistant to HIV-1 infection. HIV-1 infection in CCR5 -/- individuals, although rare, has been increasingly documented. We now report that the viral quasispecies from one such individual throughout disease is homogenous, T cell line tropic, and phenotypically syncytium inducing (SI); exclusively uses CXCR4; and replicates well in CCR5 -/- primary T cells. The recently discovered coreceptors BOB and Bonzo are not used. Although early and persistent SI variants have been described in longitudinal studies, this is the first demonstration of exclusive and persistent CXCR4 usage. With the caveat that the earliest viruses available from this subject were from approximately 4 years following primary infection, these data suggest that HIV-1 infection can be mediated and persistently maintained by viruses which exclusively utilize CXCR4. The lack of evolution toward the available minor coreceptors in this subject underscores the dominant biological roles of the major coreceptors CCR5 and CXCR4. This and two similar subjects (R. Biti, R. Ffrench, J. Young, B. Bennetts, G. Stewart, and T. Liang, Nat. Med. 3:252-253, 1997; I. Theodoreu, L. Meyer, M. Magierowska, C. Katlama, and C. Rouzioux, Lancet 349:1219-1220, 1997) showed relatively rapid CD4+ T-cell declines despite average or low initial viral RNA load. Since viruses which use CXCR4 exclusively cannot infect macrophages, these data have implications for the relative infection of the T-cell compartment versus the macrophage compartment in vivo and for the development of CCR5-based therapeutics.

Acquired Immunodeficiency Syndrome↗
PubMed1998

Steady-state plasma membrane expression of human cytomegalovirus gB is determined by the phosphorylation state of Ser900.

Human cytomegalovirus (HCMV) infection of an astrocytoma cell line (U373) or human fibroblast (HF) cells results in a differential cell distribution of the major envelope glycoprotein gB (UL55). This 906-amino-acid type I glycoprotein contains an extracellular domain with a signal sequence, a transmembrane domain, and a 135-amino-acid cytoplasmic tail with a consensus casein kinase II (CKII) site located at Ser900. Since phosphorylation of proteins in the secretory pathway is an important determinant of intracellular trafficking, the state of gB phosphorylation in U373 and HF cells was examined. Analysis of cells expressing wild-type gB and gB with site-specific mutations indicated that the glycoprotein was equally phosphorylated at a single site, Ser900, in both U373 and HF cells. To assess the effect of charge on gB surface expression in U373 cells, Ser900 was replaced with an aspartate (Asp) or alanine (Ala) residue to mimic the phosphorylated and nonphosphorylated states, respectively. Expression of the Asp but not the Ala gB mutation resulted in an increase in the steady-state expression of gB at the plasma membrane (PM) in U373 cells. In addition, treatment of U373 cells with the phosphatase inhibitor tautomycin resulted in the accumulation of gB at the PM. Interestingly, the addition of a charge at Ser900 trapped gB in a low-level cycling pathway at the PM, preventing trafficking of the protein to the trans-Golgi network or other intracellular compartments. Therefore, these results suggest that a tautomycin-sensitive phosphatase regulates cell-specific PM retrieval of gB to intracellular compartments.

Alanine↗
PubMed1998

Efficient infection of brain microvascular endothelial cells by an in vivo-selected neuroinvasive SIVmac variant.

A neuroinvasive/neuropathogenic SIV variant termed SIVmac182 was previously isolated and characterized (Watry et al, 1994). This neuroinvasive strain was derived from the uncloned strain SIVmac251 through serial animal passage of infected microglia, unlike previously reported neurovirulent strains. Importantly, the virus described here was isolated from a strain which already demonstrates limited neuroinvasiveness in vivo, through a route of inoculation which exerts selective pressure for variants in the periphery that can naturally cross the blood-brain barrier and gain access to the brain. Examination of animal tissues indicated that the neuroinvasive strain was capable of replicating in brain microvascular endothelial cells (BMEC). Therefore, we developed an in vitro model of BMEC infection in which to examine mechanisms of virus neuroinvasiveness and neuropathogenicity as well as to address mechanisms of HIV-induced dementia. Results obtained with this in vitro system indicate that growth in BMEC may predict neuroinvasiveness in vivo, and furthermore, that brain passage of virus results in the generation of neuroinvasive strains which demonstrate an increased efficiency of BMEC infection in vitro.

AIDS Dementia Complex↗
PubMed1998

Criminal and behavioral aspects of juvenile sexual homicide.

This preliminary research provides a descriptive, systematic study of juvenile sexual homicide. Fourteen incarcerated juveniles, identified through a department of corrections computer search, were assessed using a structured diagnostic interview, an author-designed clinical interview, and a review of correctional files and other available records. Five of the offenders' victims survived the homicidal attack, but their cases were nevertheless included in this study as the offenders' intent was clearly to kill their victim, and the victim's survival was merely by chance. All victims were female and all offenders were male. Their crimes typically occurred in the afternoon, and involved a low-risk victim of the same race who lived in the offender's neighborhood. The sexual component of the crime consisted of vaginal rape in over one-half of the cases. Weapons, typically a knife or bludgeon, were used in all but one case. Thirteen of these youths had a prior history of violence, and twelve had previous arrests. Chaotic, abusive backgrounds and poor adjustment in school were typical for these boys. A conduct disorder diagnosis was present in twelve of the youths, and violent sexual fantasies were experienced by one-half of the sample. The findings in this study suggest that juvenile sexual murderers comprise less than 1% of juvenile murderers, and are likely to be an emotionally and behaviorally disturbed population with serious familial, academic, and environmental vulnerabilities.

Adjustment Disorders↗
PubMed1997

Interferon-gamma and tumor necrosis factor-alpha specifically induce formation of cytomegalovirus-permissive monocyte-derived macrophages that are refractory to the antiviral activity of these cytokines.

Monocytes/macrophages are key cells in the pathogenesis of human cytomegalovirus (HCMV). Although HCMV infection in monocytes is restricted to early events of gene expression, productive infection has been demonstrated in differentiated macrophages in vitro. We examined the cellular and cytokine components that are essential for HCMV replication in Concanavalin A-stimulated monocyte-derived macrophages (MDM). By negative selection, depletion of CD8+ T lymphocytes, but not CD4+ T lymphocytes, CD19+ B cells, or CD56+ NK cells, resulted in a 60-70% reduction in the number of HCMV-infected MDM, and a 4 log decrease in virus production. Neutralization of IFN-gamma and TNF-alpha, but not IL-1, IL-2, or TGF-beta, decreased production of virus by 4 logs and 2 logs, respectively. Subsequently, addition of recombinant IFN-gamma or TNF-alpha to purified monocyte cultures was sufficient to produce HCMV-permissive MDM. While IFN-gamma and TNF-alpha possess antiviral properties, addition of these cytokines to permissive MDM cultures did not affect production of HCMV. Thus, rather than inhibiting replication of HCMV, IFN-gamma and TNF-alpha specifically induce differentiation of monocytes into HCMV-permissive MDM, which are resistant to the antiviral effects of these cytokines.

Antiviral Agents↗
PubMed1997

Tyrosine kinase inhibitors. 13. Structure-activity relationships for soluble 7-substituted 4-[(3-bromophenyl)amino]pyrido[4,3-d]pyrimidines designed as inhibitors of the tyrosine kinase activity of the epidermal growth factor receptor.

The general class of 4-(phenylamino)quinazolines are potent (some members with IC50 values << 1 nM) and selective inhibitors of the tyrosine kinase activity of the epidermal growth factor receptor (EGFR), via competitive binding at the ATP site of the enzyme, but many of the early analogues had poor aqueous solubility (<< 1 mM). A series of 7-substituted 4-[(3-bromophenyl)-amino]pyrido[4,3-d]pyrimidines, together with selected (3-methylphenyl)amino analogues, were prepared by reaction of the analogous 7-fluoro derivatives with appropriate amine nucleophiles in 2-BuOH or aqueous 1-PrOH. All of the compounds were evaluated for their ability to inhibit the tyrosine-phosphorylating action of EGF-stimulated full-length EGFR enzyme. Selected analogues were also evaluated for their inhibition of autophosphorylation of the EGF receptor in A431 human epidermoid carcinoma cells in culture and against A431 tumor xenografts in mice. Analogues bearing a wide variety of polyol, cationic, and anionic solubilizing substituents retained activity, but the most effective in terms of both increased aqueous solubility (> 40 mM) and retention of overall inhibitory activity (IC50's of 0.5-10 nM against isolated enzyme and 8-40 nM for inhibition of EGFR autophosphorylation in A431 cells) were weakly basic amine derivatives. These results are broadly consistent with a proposed model for the binding of these compounds to EGFR, in which the 6- and 7-positions of the pyridopyrimidine ring are in a largely hydrophobic binding region of considerable steric freedom, at the entrance of the adenine binding cleft. The most active cationic analogues have a weakly basic side chain where the amine moiety is three or more carbon atoms away from the nucleus. Two of the compounds (bearing weakly basic morpholinopropyl and strongly basic (dimethylamino)butyl solubilizing groups) produced in vivo tumor growth delays of 13-21 days against advanced stage A431 epidermoid xenografts in nude mice, when administered i.p. twice per day on days 7-21 posttumor implant. Treated tumors did not increase in size during therapy and resumed growth at the termination of therapy, indicating an apparent cytostatic effect for these compounds under these treatment conditions. The data suggest that continuous long-term therapy with these compounds may result in substantial tumor growth inhibition.

Animals↗
PubMed1997

Morphological and functional characterization of an in vitro blood-brain barrier model.

Cell culture models have been extensively used for studies of blood-brain barrier (BBB) function. However, several in vitro models fail to reproduce some, if not most, of the physiological and morphological properties of in situ brain microvascular endothelial cells. We have recently developed a dynamic, tridimensional BBB model where endothelial cells exposed to intraluminal flow form a barrier to ions and proteins following prolonged co-culturing with glia. We have further characterized this cell culture model to determine whether these barrier properties were due to expression of a BBB phenotype. Endothelial cells of human, bovine or rodent origin were used. When co-cultured with glia, intraluminally grown endothelial cells developed features similar to in vivo endothelial cells, including tight junctional contacts at interdigitating processes and a high transendothelial resistance. This in vitro BBB was characterized by the expression of an abluminal, ouabain-sensitive Na/K pump, and thus favored passage of potassium ions towards the lumen while preventing K+ extravasation. Similarly, the in vitro BBB prevented the passage of blood-brain barrier-impermeant drugs (such as morphine, sucrose and mannitol) while allowing extraluminal accumulation of lipophylic substances such as theophylline. Finally, expression of stereo-selective transporters for Aspartate was revealed by tracer studies. We conclude that the in vitro dynamic BBB model may become an useful tool for the studies of BBB-function and for the testing of drug passage across the brain endothelial monolayer.

Animals↗
PubMed1997

Reactivation of latent human cytomegalovirus by allogeneic stimulation of blood cells from healthy donors.

Reactivation of human cytomegalovirus (HCMV) results in severe disease in AIDS patients and immunocompromised patients receiving blood transfusions or organ or bone marrow grafts. Although the site of HCMV latency is unknown, blood cells have been implicated as a viral reservoir. In this study, we demonstrate HCMV reactivation in vitro from seven consecutive healthy donors through allogeneic stimulation of peripheral blood mononuclear cells (PBMCs). HCMV replication was detected at 17 days poststimulation, and virus was recovered after long-term culture from a macrophage expressing dendritic cell markers. Thus, these observations demonstrate that PBMCs harbor latent HCMV, which reactivates in a myeloid lineage cell upon allogeneic stimulation.

Antibodies, Viral↗
PubMed1997

Mutagenic consequences of the incorporation of 6-thioguanine into DNA.

6-Thioguanine (S6G) has been used in the treatment of acute leukemias because of its cytotoxic effect on proliferating leukemic cells. The cytotoxicity of S6G is thought to derive from its incorporation into DNA in place of guanine. The deoxyribonucleoside triphosphate of S6G, SdGTP, is a good substrate for bacterial and human DNA polymerases (Ling et al., Mol Pharmacol 40: 508-514, 1991). Since SdGTP was observed to misincorporate in place of adenine at a greater frequency than did dGTP, it appeared plausible that this analog could produce more subtle effects (mutations) due to mispairing with thymine. To assess whether such mutations occur, SdGTP was incorporated into the lacI gene of phage M13lacISaXb in reactions that omitted dGTP (-G) or dATP (-A). LacI mutation frequency was determined by beta-galactosidase colorimetric staining (inactivation of the lac repressor results in blue plaques in the absence of inducer). When a high concentration of SdGTP (24 microM) was used in the DNA polymerase reaction, phage infectivity was inhibited. When a relatively low concentration (2.4 nM) was added to the -G and -A reactions, mutagenic effects were observed. DNA sequencing of mutant progeny arising from the -G + S6G reaction revealed C-to-T base transitions and some C-to-A transversions. Similarly, the presence of SdGTP in the -A reactions led to mutants with T-to-C transitions. No insertions or deletions were observed. These data indicate that mispairing of S6G with thymine leads to mutagenic effects in this assay.

Bacterial Proteins↗
PubMed1997

Steroid hormones act directly to trigger segment-specific programmed cell death of identified motoneurons in vitro.

In larvae of the hawkmoth, Manduca sexta, accessory planta retractor (APR) motoneurons undergo a segment-specific pattern of programmed cell death at pupation. APR death is triggered hormonally by the prepupal peak of the ecdysteroid, 20-hydroxyecdysone (20-HE). Previous studies found no evidence that cellular interactions regulate the segmental pattern of APR death in vivo. To test the hypothesis that 20-HE acts directly on APRs to trigger a cell-autonomous, segment-specific pattern of death, APRs were labeled with the fluorescent dyes DiI or DiA, removed from the nervous system before the prepupal peak, and placed in low-density cell culture. Physiological levels of 20-HE triggered the same segment-specific pattern of APR death in vitro as seen in vivo, both in cultures containing a single APR and in cultures containing two APRs removed from the same donor animal. The presence or absence of contact with other cells did not influence the APRs' responses to 20-HE. The death of APRs in culture was characterized by fragmentation or rounding up of the cell body and fragmentation of the neurites. These findings suggest that intrinsic segmental identity regulates whether these motoneurons live or die when exposed to a steroid hormone during development.

Animals↗
PubMed1997

Neurocognitive impairment is an independent risk factor for death in HIV infection. San Diego HIV Neurobehavioral Research Center Group.

OBJECTIVE: To determine if mortality is increased in individuals with human immunodeficiency virus type 1 (HIV-1)-associated neurocognitive disorders less severe than frank dementia. DESIGN: A prospective cohort study; median duration of follow-up was 2.4 years. Kaplan-Meier analysis and Cox proportional hazards models were used to compare survival times according to neurocognitive classification. SETTING: University-based research unit. PARTICIPANTS: A volunteer sample of 414 individuals seropositive for HIV-1. Subjects were classified at their baseline evaluation as neuropsychologically (NP) normal or abnormal (impaired in > or = 2 NP test domains). A subgroup of NP abnormal subjects met operational criteria for HIV-associated minor cognitive motor disorder; the remaining subjects were designated NP impaired. Subjects with frank dementia were excluded. MAIN OUTCOME MEASURE: Mortality. RESULTS: At the baseline evaluation, 256 (62%) of 414 subjects were designated normal; 109 (26%). NP impaired; and 49 (12%), minor cognitive motor disorder. One hundred six participants (26%) died during follow-up. Compared with the NP normal group, the unadjusted relative risk (RR) of death for all NP abnormal subjects (minor cognitive motor disorder and NP impaired) was significantly increased (RR, 1.7; 95% confidence interval [CI], 1.2-2.6; P < .005). After adjusting for concurrently measured predictors of survival (CD4 lymphocyte counts, Centers for Disease Control and Prevention HIV disease classification, hemoglobin concentration, and serum beta 2-microglobulin) in proportional hazards models, mortality for all NP abnormal subjects remained elevated (RR, 1.8; 95% CI, 1.2-2.8; P < .01). The elevation in mortality risk for subjects with minor cognitive motor disorder was statistically significant (RR, 2.2; 95% CI, 1.2-3.8; P < .01); for NP impaired subjects it was marginally significant (RR, 1.6; 95% CI, 1.0-2.8; P = .06). CONCLUSIONS: The HIV-infected individuals with NP impairment had a higher risk of dying than those without impairment. This was particularly true for those meeting syndromic diagnostic criteria.

Adult↗
PubMed1997

Cerebrospinal fluid human immunodeficiency virus type 1 RNA levels are elevated in neurocognitively impaired individuals with acquired immunodeficiency syndrome. HIV Neurobehavioral Research Center Group.

To determine whether cerebrospinal fluid (CSF) viral burden measurements can assist in the evaluation of human immunodeficiency virus (HIV)-associated neurocognitive disorders, we quantified HIV type 1 (HIV-1) RNA in CSF. Because previous findings suggested that disease stage, lymphocytic pleocytosis, and HIV-1 RNA levels in plasma may influence CSF viral burden, these variables were examined as potential modifying factors. HIV-1 RNA levels were quantified by using a reverse transcriptase-polymerase chain reaction assay. Performance on a comprehensive neuropsychological (NP) battery was noted in 97 prospectively enrolled, HIV-infected subjects. Among subjects with acquired immunodeficiency syndrome (AIDS) (<200 CD4+ lymphocytes), NP impairment was associated with significantly higher CSF RNA levels (3.1 vs 1.8 log10 copies/ml; p = 0.02); most impaired subjects met criteria for HIV-associated dementia or minor cognitive-motor disorder. In subjects without AIDS, CSF RNA and NP impairment were unrelated. Before AIDS, CSF RNA was strongly correlated to plasma RNA and to pleocytosis, but in AIDS, CSF and plasma RNA were independent. In conclusion, we found elevated CSF HIV-1 RNA levels in NP impaired subjects with AIDS. Before AIDS, systemic viral replication, possibly through CD4+ mononuclear cell trafficking, may govern virus levels in CSF, whereas in AIDS, CD4 cell depletion may unmask a correlation between increased productive central nervous system HIV infection and clinical neurocognitive disorders.

Acquired Immunodeficiency Syndrome↗
PubMed1997

Dendritic injury is a pathological substrate for human immunodeficiency virus-related cognitive disorders. HNRC Group. The HIV Neurobehavioral Research Center.

To determine the neuropathological substrate of human immunodeficiency virus (HIV)-associated neurocognitive disorders, we examined persons with acquired immunodeficiency syndrome before their death and related their antemortem neuropsychological performance to postmortem indicators of HIV encephalitis, viral burden, and presynaptic and postsynaptic neuronal injury. Of 20 prospectively examined cases, 9 were neurocognitively normal, 5 showed neuropsychological impairment, 5 had minor cognitive/motor disorder, and 1 was demented. Degree of neurocognitive impairment was strongly related to the amount of dendritic simplification based on microtubule-associated protein 2 immunohistochemical staining, somewhat less so to a semiquantitative viral burden score based on numbers of HIV gp41-immunoreactive cells, and much less so to the presence of multinucleated giant cells or microglial nodules. It appears that even milder neurocognitive impairment reflects microneuroanatomical injury to synaptic structures.

AIDS-Related Opportunistic Infections↗
PubMed1997

Intestinal absorption of Mn-mesoporphyrin in a small bowel sac system: effect of oleic acid.

RATIONALE AND OBJECTIVES: The authors investigated the effect of oleic acid (cis-9-octadecenoic acid) (OA), a lipidic carrier, on the intestinal absorption rate and T1 relaxation time of manganese (III) mesoporphyrin (Mn-mesoporphyrin), a prototype hepatobiliary contrast agent for magnetic resonance imaging. METHODS: Mn-mesoporphyrin was formulated with OA at various concentrations. Small bowel sacs were created in 36 rats and filled with complexed and free Mn-mesoporphyrin. Intestinal absorption of Mn-mesoporphyrin was measured with spectrophotometry at 364 nm. T1 relaxation times were measured in samples of Mn-mesoporphyrin solutions, bowel wall, liver, and bile. RESULTS: Absorption rates ranged from 4.2%/cm2/h to 13%/cm2/h. Absorption was greatest (13%/cm2/h) when a combination of 1 mmol/L Mn-mesoporphyrin and 26.5 mmol/L OA was used. The T1 of bile decreased from 2,480 to 248 msec (maximum decrease) in rats that received Mn-mesoporphyrin. CONCLUSION: Mn-mesoporphyrin is absorbed from the small bowel in both the lipid-associated and free form, resulting in substantial shortening of the T1 in bile.

Animals↗
PubMed1997

Antiviral pressure exerted by HIV-1-specific cytotoxic T lymphocytes (CTLs) during primary infection demonstrated by rapid selection of CTL escape virus.

The HIV-1-specific cytotoxic T lymphocyte (CTL) response is temporally associated with the decline in viremia during primary HIV-1 infection, but definitive evidence that it is of importance in virus containment has been lacking. Here we show that in a patient whose early CTL response was focused on a highly immunodominant epitope in gp 160, there was rapid elimination of the transmitted virus strain and selection for a virus population bearing amino acid changes at a single residue within this epitope, which conferred escape from recognition by epitope-specific CTL. The magnitude (> 100-fold), kinetics (30-72 days from onset of symptoms) and genetic pathways of virus escape from CTL pressure were comparable to virus escape from antiretroviral therapy, indicating the biological significance of the CTL response in vivo. One aim of HIV-1 vaccines should thus be to elicit strong CTL responses against multiple codominant viral epitopes.

Acquired Immunodeficiency Syndrome↗
PubMed1997

HIV-1 induction of CD40 on endothelial cells promotes the outgrowth of AIDS-associated B-cell lymphomas.

Human immunodeficiency virus (HIV)-1 infection is associated with the development of aggressive extranodal B-cell non-Hodgkin's lymphomas. Using microvascular endothelial cell (MVEC)-enriched bone marrow stromal cultures, HIV infection of stromal MVECs from lymphoma patients induced the outgrowth of malignant B cells. MVECs were the only HIV-infected cells in the stroma, and purified brain MVECs also induced a phenotype supportive of neoplastic B-cell attachment and proliferation. HIV infection of MVECs stimulated surface expression of CD40 and allowed preferential induction of the vascular cell adhesion molecule VCAM-1 after CD40 triggering. B-lymphoma cells expressed the CD40 ligand (CD40L), and blocking of CD40-CD40L interactions between HIV-infected MVECs and B-lymphoma cells inhibited B-cell attachment and proliferation. These observations suggest that HIV promotes B-lymphoma cell growth through facilitating attachment of lymphoma cells to HIV-infected MVECs and represent a novel mechanism through which viruses may induce malignancies.

CD40 Antigens↗
PubMed1997

Evolutionary variants of the human immunodeficiency virus type 1 V3 region characterized by using a heteroduplex tracking assay.

Syncytium-inducing (SI) variants of human immunodeficiency virus type 1 (HIV-1) are evolutionary variants that are associated with rapid CD4+ cell loss and rapid disease progression. The heteroduplex tracking assay (HTA) was used to detect evolutionary V3 variants by amplifying the V3 sequences from viral RNA derived from 50 samples of patient plasma. For this V3-specific HTA (V3-HTA), heteroduplexes were formed between the patient V3 sequences and a probe with the subtype B consensus V3 sequence. Evolution was then measured by divergence from the consensus. The presence of evolutionary variants was correlated with SI detection data on the same samples from the MT-2 cell culture assay. Evolutionary variants were correlated with the SI phenotype in 88% of the samples, and 96% of the SI samples contained evolutionary variants. In most cases the evolutionary V3 variants represented discrete clonal outgrowths of virus. Sequence analysis of the six discordant samples that did not show this correlation indicated that three non-syncytium-inducing (NSI) samples had V3 sequences that had evolved away from the consensus sequence but not toward an SI genotype. A fourth sample showed little evolution away from the consensus but was SI, which indicates that not all SI variants require basic substitutions in V3. The other two samples had SI-like genotypes and NSI phenotypes, suggesting that V3-HTA was able to detect SI emergence in these samples in the absence of their detection in vitro. V3-HTA was also used to confirm SI variant selection in MT-2 cells and to examine the possibility of variant selection during virus culture in peripheral blood cells.

Acquired Immunodeficiency Syndrome↗