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Biomedical subjects

Robert F Miller

Publications and source records attributed to Robert F Miller.

At least 19 recordsLinked to original sources
PubMed2006

Form and function of ON-OFF amacrine cells in the amphibian retina.

ON-OFF amacrine cells were studied with whole cell recording techniques and intracellular staining methods using intact retina-eyecup preparations of the tiger salamander (Ambystoma tigrinum) and the mudpuppy (Necturus maculosus). Morphological characterization of these cells included three-dimensional reconstruction methods based on serial optical sections obtained with a confocal microscope. Some cells had their detailed morphology digitized with a computer-assisted tracing system and converted to compartmental models for computer simulations. The dendrites of ON-OFF amacrine cells have spines and numerous varicosities. Physiological recordings confirmed that ON-OFF amacrine cells generate both large- and small-amplitude impulses attributed, respectively, to somatic and dendritic generation sites. Using a multichannel model for impulse generation, computer simulations were carried out to evaluate how impulses are likely to propagate throughout these structures. We conclude that the ON-OFF amacrine cell is organized with multifocal dendritic impulse generating sites and that both dendritic and somatic impulse activity contribute to the functional repertoire of these interneurons: locally generated dendritic impulses can provide regional activation, while somatic impulse activity results in rapid activation of the entire dendritic tree.

Amacrine Cells↗
PubMed2006

Monocyte derived dendritic cells from HIV-1 infected individuals partially reconstitute CD4 T-cell responses.

OBJECTIVES: The study tests the hypothesis that monocyte derived dendritic cells from HIV-1 infected individuals are normal and can restore impaired CD4 T-cell antigen specific responses. DESIGN: Monocyte derived dendritic cells were isolated from individuals at three different stages of HIV-1 infection with a wide spectrum of viral load and CD4 T-cell counts, and from healthy volunteers. The cell surface phenotype and allogeneic stimulatory potential of these dendritic cells was documented. CD4 T-cell responses to HIV p24, tetanus toxoid and purified protein derivative were measured using either unfractionated peripheral blood mononuclear cells, or purified dendritic cell/T-cell cultures. RESULTS: Dendritic cells from all three HIV-1 infected groups did not differ from each other or from healthy volunteers in terms of cell surface phenotype or allogeneic stimulatory potential using T cells from healthy volunteers. Dendritic cells from immunosuppressed antiretroviral naive individuals enhanced the autologous recall proliferative responses both to HIV-1 p24, and third party antigens tetanus toxoid and purified protein derivative, both in terms of the proportion of responding individuals, and median proliferation. CONCLUSION: Antigen presentation by dendritic cells partially restores impaired antigen specific CD4 T-cell responses associated with HIV-1 infection. Immunization strategies which target dendritic cells may therefore offer significant advantages in the ability to stimulate HIV-specific protective immune responses.

Adult↗
PubMed2006

HIV-1 infection of mononuclear phagocytic cells: the case for bacterial innate immune deficiency in AIDS.

HIV-1 infection of mononuclear phagocytic cells, comprising monocytes, macrophages, and dendritic cells, has been the subject of extensive research over the past 20 years. The roles of mononuclear phagocytic cells in transmission of HIV-1 infection and as reservoirs of actively replicating virus have received particular attention. Experimental data have also accumulated about the effects of HIV-1 on the physiological function of mononuclear phagocytic cells, particularly their role in innate immunity to bacteria. The effects of HIV-1 on bacterial innate immune responses by mononuclear phagocytic cells are discussed here together with reports of direct interactions between HIV-encoded products and bacterial innate immune signalling pathways. These reports demonstrate mechanisms for HIV-mediated disruption of innate immune responses by mononuclear phagocytic cells that could provide novel therapeutic targets in HIV-infected patients. The clinical urgency is highlighted by greatly increased risk of invasive bacterial disease in this population, even in the era of highly active antiretroviral therapy. HIV-mediated injury to bacterial innate immunity provides an experimental paradigm that could broaden our overall understanding of innate immunity and be used to study responses to pathogens other than bacteria.

Acquired Immunodeficiency Syndrome↗
PubMed2006

Septic shock and multi-organ failure in HIV infection-'sepsis tuberculosa gravissima'.

A profoundly immunosuppressed HIV-infected man developed sepsis syndrome with multi-organ failure. A septic screen failed to identify a bacterial or fungal cause and despite empirical treatment for these pathogens the patient remained unwell. Investigations revealed disseminated tuberculosis. With specific anti-tuberculosis therapy the patient rapidly recovered. Although most cases of sepsis syndrome in HIV-infected patients are due to bacteria, tuberculosis should be added to the differential diagnosis of this presentation.

AIDS-Related Opportunistic Infections↗
PubMed2005

Performance of a T-cell-based diagnostic test for tuberculosis infection in HIV-infected individuals is independent of CD4 cell count.

The performance characteristics of the enzyme-linked immunospot assay (ELISPOT) assay (T-SPOT TB) for the diagnosis of latent tuberculosis infection in HIV-infected individuals are unknown. Given that ELISPOT enumerates Mycobacterium tuberculosis antigen-specific IFN-gamma-secreting T cells, HIV-associated immunosuppression might adversely affect test performance. However, we found that 28 out of 29 HIV-positive individuals (97%) gave evaluable test results, and performance was independent of the CD4 T-cell count. ELISPOT test performance appears to be independent of HIV-associated immunosuppression.

Adult↗
PubMed2005

Proteomic patterns of preinvasive bronchial lesions.

PURPOSE: A proteomics approach is warranted to further elucidate the molecular steps involved in lung tumor development. We asked whether we could classify preinvasive lesions of airway epithelium according to their proteomic profile. EXPERIMENTAL DESIGN: We obtained matrix-assisted laser desorption/ionization time-of-flight mass spectrometry profiles from 10-microm sections of fresh-frozen tissue samples: 25 normal lung, 29 normal bronchial epithelium, and 20 preinvasive and 36 invasive lung tumor tissue samples from 53 patients. Proteomic profiles were calibrated, binned, and normalized before analysis. We performed class comparison, class prediction, and supervised hierarchic cluster analysis. We tested a set of discriminatory features obtained in a previously published dataset to classify this independent set of normal, preinvasive, and invasive lung tissues. RESULTS: We found a specific proteomic profile that allows an overall predictive accuracy of over 90% of normal, preinvasive, and invasive lung tissues. The proteomic profiles of these tissues were distinct from each other within a disease continuum. We trained our prediction model in a previously published dataset and tested it in a new blinded test set to reach an overall 74% accuracy in classifying tumors from normal tissues. CONCLUSIONS: We found specific patterns of protein expression of the airway epithelium that accurately classify bronchial and alveolar tissue with normal histology from preinvasive bronchial lesions and from invasive lung cancer. Although further study is needed to validate this approach and to identify biomarkers of tumor development, this is a first step toward a new proteomic characterization of the human model of lung cancer tumorigenesis.

Aged↗
PubMed2005

D-Serine uptake by isolated retinas is consistent with ASCT-mediated transport.

Uptake of the neuromodulator D-serine by isolated larval tiger salamander (Ambystoma tigrinum) retinas was measured using capillary electrophoresis (CE). Excised retinas were incubated in Ringer's solution in the presence of 5 microM D-serine. The supernatant was removed after 30 min, mixed with 4-fluoro-7-nitrobenz-2-oxa-1,3-diazole (NBD-F) to fluorescently label amines and analyzed using CE. Significant D-serine uptake was observed over a period of 1.5h. This is the first observation of D-serine uptake by an intact retinal tissue. D-Serine uptake in the retina was Na(+)-dependent and blocked by l-alanine, l-threonine, and l-cysteine. This pharmacology is consistent with the sodium dependent heteroexchange expected of system ASC-type transporters.

4-Chloro-7-nitrobenzofurazan↗
PubMed2005

A computational model of the ribbon synapse.

A model of the ribbon synapse was developed to replicate both pre- and postsynaptic functions of this glutamatergic juncture. The presynaptic portion of the model is rich in anatomical and physiological detail and includes multiple release sites for each ribbon based on anatomical studies of presynaptic terminals, presynaptic voltage at the terminal, the activation of voltage-gated calcium channels and a calcium-dependent release mechanism whose rate varies as a function of the calcium concentration that is monitored at two different sites which control both an ultrafast, docked pool of vesicles and a release ready pool of tethered vesicles. The postsynaptic portion of the program models diffusion of glutamate and the physiological properties of glutamatergic neurotransmission in target cells. We demonstrate the behavior of the model using the retinal bipolar cell to ganglion cell ribbon synapse. The model was constrained by the anatomy of salamander bipolar terminals based on the ultrastructure of these synapses and presynaptic contacts were placed onto realistic ganglion cell morphology activated by a range of ribbon synapses (46-138). These inputs could excite the cell in a manner consistent with physiological observations. This model is a comprehensive, first-generation attempt to assemble our present understanding of the ribbon synapse into a domain that permits testing our understanding of this important structure. We believe that with minor modifications of this model, it can be fine tuned for other ribbon synapses.

Animals↗
PubMed2005

Isolates of Pneumocystis jirovecii from Harare show high genotypic similarity to isolates from London at the superoxide dismutase locus.

Pneumocystis jirovecii is the cause of Pneumocystis pneumonia (PCP) in humans. Isolates of P. jirovecii obtained from patients in Harare, Zimbabwe were genotyped at the superoxide dismutase locus. High genotypic similarity to isolates of P. jirovecii obtained from patients in London, UK was observed. These data provide additional support for the hypothesis that P. jirovecii is genetically indistinguishable in isolates from geographically diverse locations.

Adult↗
PubMed2005

Immune reconstitution disease associated with mycobacterial infections in HIV-infected individuals receiving antiretrovirals.

Immune reconstitution disease (IRD) in HIV-infected patients is an adverse consequence of the restoration of pathogen-specific immune responses during the initial months of highly active antiretroviral treatment (HAART). Previously subclinical infections are "unmasked" or pre-existing opportunistic infections clinically deteriorate as host immunopathological inflammatory responses are "switched on". IRD is most frequently associated with mycobacterial infections. Our literature search identified 166 published cases of IRD associated with mycobacterial infections. We review the underlying immunological mechanisms, difficulties surrounding case definition and diagnosis, the wide diversity of clinical manifestations, and treatment. The importance of screening patients for mycobacterial disease before starting HAART and the critical impact of the timing of commencement of HAART in patients receiving treatment for tuberculosis are highlighted. We also discuss the problem of IRD associated with mycobacterial diseases in developing countries where tuberculosis prevalence is high and access to HAART is currently expanding.

AIDS-Related Opportunistic Infections↗
PubMed2005

Clinical value of C-reactive protein measurements in HIV-positive patients.

The acute-phase protein C-reactive protein (CRP) is a sensitive marker of inflammation and tissue damage. We measured CRP in 109 HIV-1 antibody-positive patients admitted to hospital for investigation. In 67 patients with intercurrent infection (of whom 27 were afebrile at presentation) CRP levels were 2.2-483.5 mg/dL (normal value in the general population <3 mg/dL) and in 42 patients with alternative non-infection diagnoses CRP levels were 0.5-108.6 (median=5.9) mg/dL. Whereas in those with infections elevated CRP levels fell in response to specific therapy, values remained abnormal in those with non-infection diagnoses. CRP appears useful for diagnosis and monitoring of intercurrent infection in HIV-1 antibody-positive patients. In HIV-1 antibody-positive patients without intercurrent infection, CRP values higher than in the general population possibly reflect a sustained acute-phase response as a consequence of HIV infection per se.

AIDS-Related Opportunistic Infections↗
PubMed2004

Monitoring neurotransmitter release from isolated retinas using online microdialysis-capillary electrophoresis.

Release of neurotransmitters and other primary amine-containing analytes from intact, isolated larval salamander (Ambystoma tigrinum) retinas maintained in a 6.5-microL perfusion chamber was monitored using online microdialysis-capillary electrophoresis with laser-induced fluorescence detection (CE-LIF). Primary amines were derivatized online with o-phthaldialdehyde (OPA) and beta-mercaptoethanol. With the use of overlapping injections, the perfusate was sampled every approximately 10 s. Although separation conditions were optimized using 20 mM hydroxypropyl-gamma-cyclodextrin (HP-gamma-CD) for a number of important neuromessengers including D- and L-serine, D- and L-asparate, glutamate, GABA, serotonin, dopamine, norepinephrine, and taurine, only glutamate (0.48 +/- 0.27 microM), GABA (0.25 +/- 0.12 microM), taurine (5.5 +/- 2.1 microM), and l-serine (2.8 +/- 1.0 microM) were identified in the perfusate. Elevated levels of glutamate, GABA, and taurine were detected during stimulation with 60 mM K+. This method is the first to directly sample multiple neurotransmitters from perfused, isolated retinas and to observe changes in efflux of these neurotransmitters as a result of pharmacological stimulation.

Ambystoma↗
PubMed2004

D-Serine as a glial modulator of nerve cells.

Until the last decade, it was widely accepted that D-amino acids had no functional role in higher organisms, but that they were restricted to lower organisms, such as bacteria, where they are integrated into the proteoglycans of the cell wall. However, D-serine proved to be an effective coagonist at the "glycine-binding" site of the N-methyl-D-aspartate (NMDA) glutamate receptors, and this observation led to chemical analyses that have now revealed the presence of high levels of D-serine in the central nervous system, including many regions of the brain and retina. D-Serine has been localized to astrocytes and can be released by glutamate through stimulation of AMPA receptors. A new enzyme, serine racemase has been localized to glial cells and converts L-serine to D-serine. Degradation of D-serine takes place through D-amino acid oxidase, an enzyme once thought to metabolize D-amino acids from external sources. Although the "glycine-binding" site of NMDA receptors was initially regarded as a saturated site, evidence in many brain regions has established that this site is not saturated and is therefore modulated by interactions between glial cells and neurons. In some, but not all, studies, D-serine enhances NMDA-mediated currents; a light-evoked enhancement to NMDA currents has been reported in the retina. D-serine also plays a role in synaptic and cellular development, particularly in the cerebellum, where the normal developmental sequences underlying synapse formation onto Purkinje cells and the migration of granule cells are dependent on NMDA receptors during a time when high levels of D-serine are expressed in the Bergmann glia and other cerebellar astrocytes. D-serine must be added to the list of agents through which glial cells participate in controlling the excitability of neurons.

Animals↗
PubMed2004

Cerebral CD8+ lymphocytosis in HIV-1 infected patients with immune restoration induced by HAART.

In HIV infected persons, highly active antiretroviral therapy (HAART) has reduced both the morbidity and incidence of several disorders. Its effects on direct HIV-induced damage to the CNS remain controversial. In addition, HAART may provoke an "immune reconstitution inflammatory syndrome" (IRIS). Herein we report two patients who, despite HAART, developed a diffuse encephalopathy. Their clinical, radiological and neuropathological features are described. Immunohistochemical and PCR analyses were used to detect HIV and to exclude other viruses in brain tissue. The unusual inflammatory reaction in the brain tissue was defined by immunohistochemistry. Both patients had advanced HIV disease with low CD4 counts and high HIV "viral loads" before starting HAART. In both, HAART induced an increase in CD4 count and a marked reduction in HIV viral load, which was accompanied, in patient one, by worsening of pre-existing, and, in patient two, by development of, acute encephalopathy. At post-mortem examination, the brain of patient one showed HIV encephalitis. In addition, the brains of both patients revealed HIV-DNA by PCR, diffuse microglial hyperplasia and massive and diffuse perivascular and intraparenchymal infiltration by CD8+/CD4- lymphocytes. We suggest that the rapid immune reconstitution induced by HAART in these two patients led to a redistribution of lymphocytes into peripheral blood. This was followed by recruitment of CD8+ lymphocytes into the brain, which resulted in the diffuse infiltration described. The appearances in patient two further suggest that HIV brain infection, even without encephalitis, is sufficient to trigger this response.

Adult↗