Cell lineage analysis by intracellular injection of a tracer enzyme.

Cell lineages during development of leeches can be ascertained by injection of horseradish peroxidase as a tracer into identified cells at early stages of embryogenesis. The injected embryos continue their normal development, in the course of which horseradish peroxidase is passed on in catalytically active form to the descendants of the injected cell. The distribution of the tracer enzyme and hence of the progeny of the injected cell can then be observed at a later stage of development by staining the preparation for horseradish peroxidase.

Neuronal generation of the leech swimming movement.

The swimming movement of the leech is produced by an ensemble of bilaterally symmetric, rhythmically active pairs of motor neurons present in each segmental ganglion of the ventral nerve cord. These motor neurons innervate the longitudinal muscles in dorsal or ventral sectors of the segmental body wall. Their duty cycles are phase-locked in a manner such that the dorsal and ventral body wall sectors of any given segment undergo an antiphasic contractile rhythm and that the contractile rhythms of different segments form a rostrocaudal phase progression. This activity rhythm is imposed on the motor neurons by a central swim oscillator, of which four bilaterally symmetric pairs of interneurons present in each segmental ganglion appear to constitute the major component. These interneurons are linked intra- and intersegmentally via inhibitory connections to form a segmentally iterated and inter-segmentally concatenated cyclic neuronal network. The network appears to owe its oscillatory activity pattern to the mechanism of recurrent cyclic inhibition.

The dilemma of science and morals.

The conflicts between science and morals which still continue to arise despite the apparent hegemony of atheistic scientism over traditional Judeo-Christianity in the twentieth century reflect a basic contradiction in the metaphysical foundation of Western lives. As was set forth by Machiavelli, the contradiction inherent in Western ethics is that it is based on the simultaneous belief in both objectively valid moral truths and purely relative values of communal purpose. The achievements of twentieth century science have intensified these contradictions. Modern physics has put in question the validity of its own metaphysical basis, namely the belief in Natural Law, and modern biology has been unable to come to terms with the Cartesian dualism of body and soul. By contrast, Chinese lives are comparatively free of these contradictions, being founded on the philosophies of Confucianism and Taoism, to which the concepts of objectively valid truth or Natural Law are foreign. Recent developments in Western attitudes regarding science and morals can be interpreted as a movement away from the traditional belief in absolute truths towards a Chinese relativism.

Expression of CD11/CD18 and ICAM-1 on monocytes and lymphocytes of HIV-1-infected individuals.

The role of the leukocyte integrins in the infection of cells by HIV-1 or in the progression of AIDS-related disease is of continuing interest. Using a dual-labeling flow cytometric method, we determined the level of expression of the leukocyte integrins and intercellular adhesion molecule 1 (ICAM-1) on monocytes and lymphocytes from HIV-1-infected and uninfected individuals. The mean fluorescence of CD18 on lymphocytes and CD11c on monocytes of HIV-1-infected subjects was significantly higher than for the control group (P = .008 and .014, respectively). There was a trend toward higher fluorescence of CD11a on lymphocytes from HIV-1-infected subjects compared with cells from the control group (P = .089). Integrin expression on lymphocytes or monocytes from HIV-infected individuals did not correlate with their CD4 lymphocyte number. However, the mean fluorescence associated with ICAM-1 on monocytes and CD11a and CD18 expression on lymphocytes was related to clinical stage of disease.

Macrophage tropism: fact or fiction?

Primary HIV-1 isolates can be distinguished by phenotypic qualities such as the ability to productively infect cells of established CD4-positive lines and to induce syncytia in MT-2 cells. Such viral phenotypes have also been reported to confer host cell specificity. It is perceived that primary isolates with the syncytium-inducing phenotype (SI or rapid/high) are T cell tropic and are therefore unable to infect primary cells of the monocyte/macrophage lineage. However, we have consistently found that these isolates are as capable of establishing infection in monocyte-derived macrophages (MDM) as the monocytotropic, non-syncytium-inducing variants (NSI or slow/low). It is known that differentiation, activation, and proliferation of human monocytes are affected by both isolation methods and culture conditions. Therefore, to test whether our inability to discriminate macrophage tropic HIV-1 isolates could be explained by differences in culturing techniques, we isolated monocytes by elutriation or short-term adherence and allowed the cells to mature and differentiate in either the presence or absence of autologous lymphocytes. After removal of nonadherent cells, MDM were infected with a panel of SI and NSI primary HIV-1 isolates. MDM were susceptible to infection by the SI as well as the NSI isolates, regardless of whether or not the cells were allowed to mature in the presence of autologous lymphocytes. However, MDM matured in the presence of autologous lymphocytes replicated HIV-1 isolates (both NSI and SI) to a higher titre than MDM matured in the absence of lymphocytes. In light of these findings and recently published reports on HIV-1 phenotype and chemokine receptor usage we believe that the term macrophage-tropic strains of HIV-1 is no longer appropriate.

Developmental cell lineage.

Studies of the role of cell lineage in development began in the 1870s, fell into decline in the first half of the 20th century, and were revived in the 1960s. This revival was attended by the introduction of new and powerful analytical techniques. Cell lineage can be inferred to have a causative role in developmental cell fate in embryos in which induced changes in cell division pattern lead to changes in cell fate. Such a causative role of cell lineage is suggested also by cases where homologous cell types characteristic of symmetrical and longitudinally metameric body plan arise via homologous cell lineages. The developmental pathways of commitment to particular cell fates proceed according to a mixed typologic and topographic hierarchy, which appears to reflect an evolutionary compromise between maximizing the ease of ordering the spatial distribution of determinants of commitment and minimizing the need for migration of differentially committed embryonic cells.

Deregulation of the CD95/CD95L system in lymphocytes from patients with primary acute HIV infection.

OBJECTIVE: To analyze the role of CD95/CD95 ligand (CD95L) expression and functionality in peripheral blood lymphocytes (PBL) during primary, acute HIV syndrome (AHS) and in the subsequent period. PATIENTS: Twelve patients were studied during the acute phase of the viral infection and most were followed for some months. METHODS: Cell culture and cytotoxicity assays based upon 51Cr release and flow cytometry were used to evaluate cell killing via CD95 molecule, flow cytometry to assess surface antigens, enzyme-linked immunosorbent assay (ELISA) for the determination of soluble CD95 and CD95L plasma levels, quantitative competitive (QC) reverse transcription polymerase chain reaction (RT-PCR) with an original RNA competitor for the analysis of CD95L mRNA expression and QC RT-PCR for determining plasma viral load. RESULTS: The analysis of PBL during this phase revealed that almost all cells, including CD8 T cells with a virgin phenotype, B lymphocytes and natural killer cells displayed CD95 molecules on the plasma membrane. Activation of CD95 on the surface of isolated lymphocytes by anti-CD95 monoclonal antibodies or binding to CD95L induced rapid apoptosis. However, CD95L mRNA was not expressed in PBL from these patients and was poorly inducible. Soluble CD95 was found in the plasma of all patients, but only in a few at high levels, even some months after seroconversion. In contrast, soluble CD95L was detected in only one patient, this occurring after the symptomatic period. For 10 of the 12 patients, expression of CD95 on the cell membrane or in the plasma did not correlate with the plasma viral load, which varied widely from patient to patient. Further, plasma levels of soluble CD95 were not altered by decreased lymphocyte activation or by efficient antiretroviral therapy. CONCLUSIONS: In patients experiencing an acute, primary HIV infection, a prolonged deregulation of the CD95/CD95L system may exist, which is probably not entirely related to virus production but may contribute to the pathogenesis of the disease. The hypothesis can be put forward that a complex balance exists between proapoptotic events (increase in CD95 expression), probably triggered by the host as a method to limit viral production, and antiapoptotic events (decrease in CD95L expression) probably triggered by the virus as a way to increase its production and survival.

Correlates of apoptosis of CD4+ and CD8+ T cells in tonsillar tissue in HIV type 1 infection.

The immunopathogenesis of human immunodeficiency virus type 1 (HIV-1) infection has been associated with increased death by apoptosis of T cell subsets. In the present study, we have examined correlates of apoptosis of CD4+, CD8S+CD28+, and CD8+CD28- T cells in tonsillar lymphoid tissue in persons with HIV-1. Single-cell suspensions of tonsillar lymphocytes were analyzed by flow cytometry to determine the fraction of cells showing typical characteristics of apoptosis as well as the expression of activation markers within the live and the apoptotic cell populations. The proportion of cells carrying infectious provirus was quantified by limiting dilution analysis. Compared with uninfected controls, apoptosis of both CD4+ and CD8+ T cells was enhanced in HIV-1 infection and was higher among CD8+ than among CD4+ T cells. Apoptosis of CD28-cells was more prevalent than apoptosis of CD28+ cells for both CD4+ and CD8+ T cells. Occurrence of apoptosis of CD4+ T cells correlated with provirus levels and proportional expression of the activation marker HLA-DR. Apoptosis of CD8+CD28+ cells correlated with expression of the activation markers CD69 and HLA-DR while apoptosis within CD8+CD28- cells did not correlate with any of the studied parameters. Although apoptosis was much more prevalent among CD8+ than CD4+ T cells, CD8+ T cells still accumulated in tonsillar lymphoid tissue in persons with HIV-1. Our data may be interpreted to suggest that apoptosis of CD4+, CD8+CD28+, and CD8+CD28- cells in tonsillar tissue is regulated by different mechanisms and the results are of importance to our understanding of the immunopathogenesis of HIV-1 infection.

Effects of monocyte purification and culture on integrin expression.

Selective alterations in the surface expression of members of the LeuCAM (leukocyte cell adhesion molecule) family of integrins occur during in vitro culture of human monocytes. Such changes may relate in part to cellular maturation, but also to activation following purification and culture of monocytes. In this paper, we examined the effects of monocyte isolation, adherence during culture and endotoxin exposure on the expression of these molecules and the ligand for LFA-1, ICAM-1 (CD54). Expressions of CD11b, CD18 and CD54, but not CD11a or CD11c, were higher on monocytes freshly isolated by density gradient separation and plastic adherence as compared with cells labelled directly in whole blood. However, the surface expression of the LeuCAMs and CD54 on cultured monocytes was not affected by short-term adherence to plastic for 2 h, as determined by comparisons of their expression on adherence-isolated and elutriated monocytes. In contrast, prolonged adhesion of monocytes for up to 21 days in culture altered expression of CD11a without affecting that of the other LeuCAMs or CD54. Expression of CD11a decreased more rapidly on adherence-maintained cells as compared with suspension-cultured cells. Our results show that cellular manipulations required for in vitro studies of monocyte/macrophages may alter expression of the LeuCAMs.

Isolation of human tonsillar dendritic cells.

Tonsillectomy remains a frequently performed operation in developed countries ensuring that tonsils are the most readily available source of human lymphoid tissue and an easily accessible source of dendritic cells (DC). Tonsil lymphoid tissue also provides a source of the different DC that are resident within the B- and T-cell microenvironments. Although an alternative model for follicular dendritic cell (FDC) ontogeny has been proposed (1) the FDC within tonsil B cell areas probably develop in situ from mesenchymal precursors (2). Whatever their origin, the phenotype and function of FDC (3) seem to be unrelated to the bone-marrow-derived DC that are the subject of these protocols. The precise relationship between the distinct sub-populations of the bone-marrow-derived DC within the tonsil is still not clear (see ref. 4 for review).

Multichannel audio monitor for detecting electrical signals.

The multichannel audio monitor (MUCAM) permits the simultaneous auditory monitoring of concurrent trains of electrical signals generated by as many as eight different sources. The basic working principle of this device is the modulation of the amplitude of a given pure tone by the incoming signals of each input channel. The MUCAM thus converts a complex, multichannel, temporal signal sequence into a musical melody suitable for instant, subliminal pattern analysis by the human ear. Neurophysiological experiments requiring multi-electrode recordings have provided one useful application of the MUCAM.

Prevalence of antibody to human T-lymphotropic virus type-1 (HLTV-1) in Australian aborigines, and detection in Indonesian sera.

The first finding of antibody to human T-lymphotropic virus type 1 in Australia, specifically in Australian Aborigines, is reported. The overall results suggest that this is a new area to be added to the known endemic areas for this virus. Antibody prevalence in each of two widely separated areas was found to be approximately 16% in 1977, and in one of these areas this had increased to approximately 34% in 1984/86. In this area no antibody to this virus was detected in children under 4 years of age.

PIP: The 1st finding of antibody-to-human T-lymphotropic virus type 1 in Australia, specifically in Australian Aborigines, is reported here. Overall results suggest that this is a new area to be added to those known endemic areas for this virus. Antibody prevalence in each of 2 widely separated areas were found to be approximately 16% in 1977, and in 1 of these areas, this increased to approximately 34% in 1984-86. No antibody to this virus was detected in this area in children under 4 years of age.