Biomedical subjects

C L Miller

Publications and source records attributed to C L Miller.

At least 37 records · Page 2Linked to original sources

PubMed1997

Differential ability of flt3-ligand, interleukin-11, and Steel factor to support the generation of B cell progenitors and myeloid cells from primitive murine fetal liver cells.

A variety of factors produced by stromal fibroblasts, including Flt3-ligand (FL), interleukin-11 (IL-11), Steel factor (SF), and IL-7, have been implicated in stimulating the production of pre-B cells and myeloid cells from primitive hematopoietic precursors. To investigate their relative roles in this process, either as single-acting or synergistic agents, we compared the yield and types of cells produced after 2 weeks from small numbers of Sca-1+ Lin- (i.e., B220-, Ly-1-, Gr-1-, and Ter-119-) day 14.5 murine fetal liver cells placed in stromal cell-free cultures containing all possible combinations of FL, SF, IL-7, and IL-11. None of these factors alone supported the production (or survival) of any cells beyond 1 week: only pairs of factors consisting of either FL or SF plus either IL-11 or IL-7 were effective in this regard, with FL plus IL-11 being the most potent pair (approximately 7 x 10(4) cells obtained per 100 Sca-1+ Lin- input cells). The maximum numbers of cells were produced in the presence of FL, IL-11, and IL-7: these included both B220+ and Mac-1+/Gr-1+ cells (approximately 10(6) and approximately 2 x 10(5), respectively, per 100 Sca-1+ Lin- input cells). Both of these lineages were also obtained with each of the other possible three-factor combinations, albeit with variable effectiveness. Omission of either FL or IL-7 caused the greatest reduction in the yield of B220+ cells (approximately 130-fold and approximately 80-fold, respectively). Omission of IL-11 and, to a lesser extent, FL caused the greatest reduction in the yield of Mac-1+/Gr-1+ cells (approximately 90-fold and approximately 3-fold, respectively). When fetal calf serum was replaced with a defined serum substitute, the out put of B220+ cells remained the same but myelopoiesis was consistently enhanced (approximately 5- to 20-fold). These findings support a model involving factor redundancy in the extracellular signals required to stimulate the production and amplification of both lymphoid and myeloid cells from early Sca-1+ Lin- cells. They also reveal quantitative differences in the abilities of different competent factor combinations to promote this process, which may be further modulated by the presence of undefined serum components.

Animals↗

PubMed1996

The repopulation potential of fetal liver hematopoietic stem cells in mice exceeds that of their liver adult bone marrow counterparts.

Varying, limiting numbers of unseparated or purified cells (Ly-5.1), either from 14.5-day-old fetal liver (FL) or from adult bone marrow (BM) were coinjected with 10(5) unseparated BM cells (Ly-5.2) into lethally irradiated adult C57B1/6 recipients (Ly-5.2). The kinetics of donor cell repopulation of the lymphoid and myeloid compartments by Ly-5.1+ donor hematopoietic stem cells (ie, competitive repopulation units [CRU]) were monitored at various time points after the transplantation by Ly-5 analysis of the peripheral white blood cells (WBC). Recipients that had received on average less than 2 adult BM or FL CRU did not show a significant difference in the level of donor-reconstitution when analyzed 4 weeks after the transplantation, However, at 8 and 16 weeks, the FL recipients showed a significantly higher percentage of donor-derived nucleated peripheral blood cells than did the recipients of adult BM cells. Analysis of individual mice showed that approximately 80% of the recipients of FL CRU showed an increase in mature WBC output between 4 and 8 weeks after transplantation, whereas this occurred in less than 40% in the recipients of adult BM cells. In addition to this effect on mature cell output, the cellularity of the reconstituted BM was significantly higher in recipients of FL CRU than in recipients of adult BM CRU, even at 7 to 9 months after transplantation, which is consistent with an increased clonal expansion of FL CRU. When marrow cells from primary recipients of FL CRU were injected into secondary recipients, a significantly higher percentage of these mice showed donor-reconstitution of their lymphoid and myeloid compartments (P < .01) and to a greater extent (P < .008) as compared with mice that had received marrow cells from primary recipients of similar numbers of adult BM CRU. Taken together, these results show that individual FL CRU exhibit a greater proliferative activity in vivo than similar cells from adult BM that is accompanied by a greater production of daughter CRU.

Animals↗

PubMed1996

Schistosoma mansoni: L-glutamate-induced contractions in isolated muscle fibers; evidence for a glutamate transporter.

Schistosoma mansoni muscle fibers contract in response to L-glutamate in a dose-dependent manner (10(-6)-10(-3) M). L-aspartate and D-aspartate are likewise effective in eliciting contraction of the fibers. Mammalian glutamate receptor agonists produce little or no contraction at concentrations as high as 1 mM. In addition, common glutamate receptor antagonists do not inhibit the contraction induced by L-glutamate. However, amino acids known to be substrates for the high-affinity glutamate transporter elicit contraction of the muscle fibers. These results suggests that there is a high-affinity glutamate transporter on the muscle fibers which, because of its electrogenic nature, is causing depolarization and contraction. This is supported by the evidence that contraction induced by L-glutamate is dependent on extracellular Ca2+ and is blocked by nicardipine (10 microM). [3H]L-glutamate is taken up in a dose-dependent manner by the muscle fiber preparation. This uptake is also time- and temperature-dependent. Both the L-glutamate-induced contractile response and [3H]L-glutamate uptake are Na(+)-dependent and can be blocked by specific inhibitors of the high-affinity transporter. This experimental evidence supports the hypothesis that there is a Na(+)-dependent high-affinity glutamate transporter on the schistosome muscle membrane.

ATP-Binding Cassette Transporters↗

PubMed1996

Regulation of Epstein-Barr virus latency by latent membrane protein 2.

Like other herpesviruses, Epstein-Barr virus persists in its host through its ability to establish a latent infection that periodically reactivates. Latent membrane protein 2A (LMP2A) regulates reactivation from latency by interfering with normal B cell signal transduction processes, and may define a new class of regulators of herpesvirus latency.

Amino Acid Sequence↗

PubMed1996

Neuromuscular physiology and pharmacology of parasitic flatworms.

The trematode and cestode flatworms include numerous parasitic forms of major medical and economic importance. A better knowledge of the neuromuscular physiology of these animals could lead to development of new control measures against these parasites. Since these animals are near the stem from which all other animals have evolved, better knowledge of these animals could also yield valuable information about the early evolution of nerve and muscle systems in the animal kingdom. This review focuses on what is known about the characteristics of the somatic muscle in these animals. The anatomy of the muscles is described along with a review of current information about their electrophysiology, including descriptions of the ion channels present. Also included is a summary of recently acquired data concerning the nature of serotonin, peptide, acetylcholine and glutamate receptors on the membranes of the muscles.

Animals↗

PubMed1996

EBNA-2 and EBNA-3C extensively and mutually exclusively associate with RBPJkappa in Epstein-Barr virus-transformed B lymphocytes.

Although genetic and biochemical data indicate that the cell protein RBPJkappa is a mediator of EBNA-2 and EBNA-3C effects on transcriptional regulatory elements, the extent of association of these Epstein-Barr virus nuclear proteins with RBPJkappa in transformed B lymphocytes has not been determined. We now report that most of the EBNA-2 and at least 20% of the EBNA-3C coimmunoprecipitated with RBPJkappa from extracts of transformed B lymphocytes that contained most of the cellular EBNA-2 and EBNA3C. Both proteins are associated preferentially with the smaller of the two RBPJkappa isoforms. EBNA-2-RBPJkappa complexes do not contain EBNA-3C, and EBNA-3C-RBPJkappa complexes do not contain EBNA-2. Although EBNA-2 and EBNA-3C are extensively associated with RBPJkappa, a fraction of RBPJkappa appears to be free of EBNAs after repeated immunoprecipitations with anti-EBNA, Epstein-Barr virus-immune, human antibody. Promoters with RBPJkappa sites in their regulatory elements are likely to be differentially regulated by these RBPJkappa-EBNA-2 and RBPJkappa-EBNA-3 complexes.

Antigens, Viral↗

PubMed1996

Severity of medical and neurologic complications as a determinant of neurodevelopmental outcome at 6 and 12 months in very low birth weight infants.

Very low birth weight (n = 154) and term infants (n = 119) had neurologic and developmental assessment at 6 and 12 months of age. Preterm infants with severe neonatal complications were considered to be at high risk, and those with milder complications were considered to be at low risk, for neurodevelopmental abnormality. Compared to term infants, high- and low-risk infants had abnormalities at 6 months in total neurologic score, cranial nerves, motor tone, motor coordination, and reflexes (P < .001). At 12 months, all groups had improved. However, high-risk infants had persistent abnormalities in the same subcategories (P < .001), whereas low-risk infants differed from term infants only in motor tone (P < .001). Bayley developmental scores were different for all groups at 6 months (P < .001), but at 12 months only high-risk infants differed from term infants (P < .01). These results demonstrate improvement in neurologic and developmental scores over time in very low birth weight infants. The degree of neurodevelopmental abnormality and improvement over time is related to severity of neonatal complications in preterm infants.

Age Factors↗

PubMed1996

A comparison of long-term repopulating hematopoietic stem cells in fetal liver and adult bone marrow from the mouse.

Previous studies have shown that stem cells able to competitively reconstitute the hematopoietic system of lethally irradiated mice (competitive repopulating units [CRU]) can be obtained in highly purified form from adult mouse bone marrow (BM) by the isolation of cells with a Sca-1+Lin-WGA+ phenotype. We now report on the phenotypic characteristics of CRU from day-14.5 murine fetal liver (FL). Our results confirm previous reports of similarities between the two CRU populations but also reveal a few striking differences. Both were found to express the Sca-1 antigen (SCA-1+ and surface molecules that bind wheat germ agglutinin (WGA+), and both show an absence or low expression of a number of markers characteristic of mature hematopoietic cells: B220, Gr-1,ly-1 and Ter119 (together termed Lin*-). Limiting dilution analysis of recipients transplanted with purified Sca-1+Lin*- FL cells with intermediate forward- and side-scatter properties showed that the frequency of CRU in this FL subpopulation was one in 39 cells. This represents an enrichment of approximately 450-fold over the labeled but unseparated FL starting population (one in 17,300 total FL cells). These FL CRU also resembled their counterparts in adult BM in that they expressed high levels of MHC class I and CD43 and intermediate levels of heat-stable antigen (HSA) and c-kit and did not express, or expressed at a low level, Thy-1.2, CD71, and the antigen recognized by the Fall-3 monoclonal antibody (mAb). In contrast, a high percentage of the Sca-1+Lin*- cells isolated from 14.5-day-old FL stained with the AA4.1, anti-Mac-1, and the anti-CD45RB mAbs and retained Rhodamine 123 (Rh123(bright)), whereas the Sca-1+Lin-WGA+ CRU-containing fraction of adult BM cells was found to be AA4.1-, Mac-1-, CD45RB-, and Rh123(dull). These differences in phenotype between CRU in FL and adult BM indicate changes that occur during ontogeny in cells that are similar with respect to their totipotentiality and long-term repopulating potential and complement parallel observations of functional differences between these two populations of CRU.

Animals↗

PubMed1996

Studies of W mutant mice provide evidence for alternate mechanisms capable of activating hematopoietic stem cells.

Previous studies have suggested that Steel factor (SF) can influence the behavior of many types of hematopoietic progenitor cells both in vivo and in vitro, although whether these may include the most primitive populations of totipotent repopulating cells remains controversial. To approach this question, we measured the number of Sca1+Lin-WGA+ cells, the number of cells with demonstrable myeloid (long-term culture-initiating cell [LTC-IC]) or both myeloid and lymphoid (LTC-IC(ML)) potential in 4- to 5-week-old long-term cultures containing irradiated primary marrow feeder layers, and the number of multilineage long-term in vivo repopulating cells (competitive repopulating unit [CRU]) present in the marrow of W42/+ or W41/W41 mice compared to +/+ controls. There was no significant effect of either of these W mutations on the number of Sca1+Lin-WGA+ cells and, in W41/W41 mice, neither LTC-IC nor LTC-IC(ML) populations appeared to be affected. On the other hand, although W41/W41 and W42/+ cells could both be detected in the in vivo CRU assay, their numbers were markedly reduced (17- and seven-fold, respectively) in spite of the fact that both of these W mutant genotypes contained near normal numbers of day-9 and -12 colony-forming units-spleen (CFU-S). In vitro quantitation of erythroid (burst-forming units-erythroid [BFU-E]), granulopoietic (CFU-granulocyte/macrophage [CFU-GM]), multilineage (CFU-granulocyte/erythrocyte/monocyte/macrophage [CFU-GEMM]), and pre-B clonogenic progenitors (CFU-pre-B) also revealed no differences in the numbers (or proliferative potential) of any of these cells when W41/W41 or W42/+ and normal mice were compared, although day 3 BFU-E from both types of W mutant mice showed no response to the typical enhancing effect exerted by SF on their +/+ counterparts. Taken together, these findings are consistent with the view that SF activation of c-kit receptor-induced signaling events is not a rate-limiting mechanism controlling red blood cell production during normal development until hematopoietic cells differentiate beyond the day-3 BFU-E stage. Nevertheless, normal hematopoietic stem cells do appear to be responsive to SF, since their W mutant counterparts display a disadvantage in the in vivo setting which is exaggerated under conditions of hematopoietic regeneration. On the other hand, alternative mechanisms also appear to contribute to the regulation of hematopoietic stem cell numbers in vivo and to their detection as LTC-IC in vitro.

Anemia↗

PubMed1995

Priming of locomotor initiation by electrical stimulation in the hypothalamus and preoptic region in the anesthetized rat.

Electrical stimulation at a locomotor site can prime (i.e., shorten the latency to initiate) stepping elicited by subsequent stimulation of the same or a different site. We tested for the priming effect in representative sites along the medial forebrain bundle, and determined if its magnitude showed regional differences. Rats (n = 20) were anesthetized with Nembutal and held in a stereotaxic apparatus over a wheel. Stepping was detected by accelerometers attached to the hindlimbs. Priming and test trains of stimulation (0.5-ms cathodal pulses, 50 Hz, 25-75 microA, 7-9-s train duration) separated by 20 s were delivered every 90 s. When the priming and test stimulations were applied to the same site, the priming effects were similar along the entire extent of the medial forebrain bundle. When the priming and test sites were different, the priming effect depended on their relative positions. Anterior stimulation primed posterior sites at magnitude comparable to those produced by stimulating the same posterior site. Posterior stimulation primed anterior sites at a level half of that produced by stimulation of the same anterior site. This pattern was found for priming and test sites that were ipsilateral and contralateral. Priming is a general and robust phenomenon with properties that may be useful for studying locomotor initiation pathways.

Anesthesia↗

PubMed1995

The activity of hippocampal interneurons and pyramidal cells during the response of the hippocampus to repeated auditory stimuli.

Interneurons and pyramidal cells were identified electrophysiologically in the hippocampus of anesthetized rats. Paired tones were presented 500 ms apart, and the resulting neuronal response was compared to differences in the amplitude of an auditory evoked potential (N40) elicited by each of the tones. Generally, the N40 elicited in response to the second tone is less than the response to the first. Pyramidal cells exhibited their most pronounced activation within 40 ms after the first tone. The post-stimulus discharge activation of interneurons was less than the pyramidal cells, but activation of different interneurons occurred at various times throughout the interval between the two tones. The presence or absence of suppression of N40 amplitude in the paired stimulus paradigm correlated with differences in the auditory response of both the interneurons and pyramidal cells. The activity of interneurons in relationship to gating of hippocampal auditory response is discussed.

Acoustic Stimulation↗

PubMed1995

Integral membrane protein 2 of Epstein-Barr virus regulates reactivation from latency through dominant negative effects on protein-tyrosine kinases.

An Epstein-Barr virus-encoded protein, LMP2, blocks the effects of surface immunoglobulin (slg) cross-linking on calcium mobilization and on lytic reactivation of EBV in latently infected and growth-transformed primary human B lymphocytes. In wild-type EBV-transformed cells, LMP2 is constitutively tyrosine phosphorylated and is associated with Lyn and Syk protein-tyrosine kinases (PTKs). Baseline Lyn PTK activity is substantially reduced, and slg cross-linking fails to activate Lyn, Syk, Pl3-K, PLC gamma 2, Vav, Shc, and MAPK. Syk, Pl3-K, PLC gamma 2, and Vav are constitutively tyrosine phosphorylated, and their tyrosine phosphorylation does not change following slg cross-linking. In contrast, cross-linking slg on cells transformed by LMP2 null mutant EBV recombinants triggers the same protein tyrosine kinase cascade as in noninfected B lymphocytes. These data are consistent with a model in which LMP2 is a constitutive dominant negative modulator of slg receptor signaling through its effects on Lyn, Syk, or regulators of these kinases.

Antigens, Viral↗

PubMed1995

Long-term potentiation disrupts auditory gating in the rat hippocampus.

The consequence of long-term potentiation (LTP) of hippocampal commissural inputs was investigated in an auditory gating paradigm. Auditory evoked potentials (AEPs) were recorded in the CA3b region of the hippocampus of rats anesthesitized with chloral hydrate. Two tones were delivered 0.5 sec apart; in this paradigm, the second AEP is diminished compared to the first. Electrical stimulation was applied to hippocampal commissural fibers to generate field potentials and population spikes which were recorded at the same site as the AEPs. LTP of the commissural input (initiated by three trains of 250 Hz/1 sec stimulation) was associated with changes in the AEPs: on average, the response to the first tone decreased and the response to the second tone increased, resulting in the disruption of auditory gating. When high-frequency stimulation of the commissural input failed to result in LTP, no effect on the AEPs was seen. If 3-(2-carboxypiperazin-4-yl)-propyl-L-phosphonic acid (CPP; 6 mg/kg, i.p.), an antagonist to the NMDA subclass of glutamate receptors, was administered prior to high-frequency stimulation, LTP induction was blocked and AEPs were not affected. Finally, reversal of LTP, achieved by high-frequency stimulation of CA3 input that was heterosynaptic to the particular commissural fibers at which the LTP was originally generated, caused disrupted auditory gating to return to normal. A model of reciprocal LTP and heterosynaptic depression of commissural and auditory input pathways is proposed to explain these findings.

Acoustic Stimulation↗

PubMed1995

Nitric oxide therapy for persistent pulmonary hypertension of the newborn.

Nitric oxide has recently been introduced to the world of neonatal pulmonary medicine. The discovery of endogenous nitric oxide production has stimulated extensive research into vascular biology to find the relation of nitric oxide to the transition of circulatory patterns at birth. Persistent pulmonary hypertension of the newborn (PPHN) has long been recognized as a neonatal complication to a variety of disorders. Traditional treatments for PPHN have been associated with adverse effects. Nitric oxide is considered the only selective pulmonary vasodilator to date. Nitric oxide therapy does not produce systemic hypotension, often associated with the use of intravenous vasodilators. This new therapy may possibly reduce or eliminate the need for invasive life-saving procedures such as extracorporeal membrane oxygenation. This article discusses in great detail nitric oxide chemistry and physiology and the procedure for nitric oxide delivery by inhalation and reviews the results of recent research.

Humans↗

PubMed1995

Nonlinear effects of radiation dose on donor-cell reconstitution by limited numbers of purified stem cells.

Despite the increasing use of bone marrow transplantation (BMT) as a treatment for a wide variety of diseases, the numbers and types of cells required for both rapid and sustained recovery of hematopoiesis are not well defined. To investigate further the potential of transplants consisting of highly enriched populations of long-term repopulating cells, we transplanted a series of Ly-5.2 mice given various doses (750, 800, 850, 900, or 950 cGy) of total-body irradiation (TBI) with 30 or 90 Sca-1+Lin-WGA+ marrow cells isolated from congenic Ly-5.1 donors. As expected, mature progeny derived from these cells, belonging to both myeloid and lymphoid compartments, could be detected with increasing case in recipients given radiation doses from 750 to 900 cGy TBI. Surprisingly, expression of this potential was significantly reduced in mice that had received 950 cGy TBI. This contrasts with the capacity of the same number of purified Sca-1+Lin-WGA+ cells to generate readily detectable progeny in 950 cGy treated mice given a simultaneous transplant of 10(5) normal marrow cells or 2 x 10(5) serially passaged marrow cells. We suggest that this variable behavior of purified stem cells in differently treated recipients may reflect radiation dose-dependent differences in the types or levels of expression of factors that regulate transplanted stem cell proliferation and differentiation in vivo and that above a certain threshold radiation dose, this may result in an irreversible loss of long-term reconstituting potential. Regardless of the nature of the underlying mechanism, this study shows that the extent of donor repopulation after BMT can be a function not only of the number of stem cells transplanted but also of the conditioning of the recipient and whether other cell types are also injected.

Animals↗

PubMed1994

An integral membrane protein (LMP2) blocks reactivation of Epstein-Barr virus from latency following surface immunoglobulin crosslinking.

The role of latent membrane protein 2 (LMP2) in Epstein-Barr virus (EBV) infection was evaluated by using latently infected primary B lymphocytes that had been growth transformed by wild-type or specifically mutated EBV recombinants. LMP2 null mutant recombinant EBV-infected cells were similar to normal B lymphocytes in their rapid increase in intracellular free calcium after surface immunoglobulin crosslinking. These cells also became more permissive for lytic EBV replication. In sharp contrast, wild-type control infected cells had little or no increase in intracellular free calcium or in permissivity for EBV replication. The block to surface immunoglobulin crosslinking-induced permissivity in cells expressing wild-type LMP2 could be bypassed by raising intracellular free calcium levels with an ionophore and by activating protein kinase C with phorbol 12-myristate 13-acetate. LMP2A, not LMP2B, mediates this effect on calcium mobilization. Genetic and biochemical data are consistent with these effects being due to the interaction of the LMP2A N-terminal cytoplasmic domain with B lymphocyte src family tyrosine kinases.

Amino Acid Sequence↗

PubMed1994

Monitoring protein kinase and phosphatase reactions with matrix-assisted laser desorption/ionization mass spectrometry and capillary zone electrophoresis: comparison of the detection efficiency of peptide-phosphopeptide mixtures.

Matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) and capillary zone electrophoresis (CZE) were evaluated for monitoring protein phosphatase and kinase reactions in vitro. Varying concentrations of peptide C (YIHLEKKYVRRDSG), peptide S (YLIEDNEYTARQGA) and kemptide (LARRSALG) mixed with their corresponding phosphorylated peptides, pC, pS and pkemptide, were analyzed. Comparison between the two techniques indicated that MALDI MS was less quantitative than CZE, showing a bias towards detection of the unphosphorylated peptide S and kemptide. In terms of sensitivity, the MALDI MS and CZE techniques are comparable. Protein kinase A phosphorylation of kemptide was monitored with both MALDI MS and CZE, whereas alkaline phosphatase dephosphorylation of pC could only be monitored with MALDI MS. The absence of inhibition with phosphatase or kinase buffers is a significant advantage of MALDI MS. In contrast to CZE, the MALDI spectra allow identification of the species analyzed by virtue of their mass. The results obtained emphasize the advantage of monitoring enzymatic reactions in buffer solutions using MALDI MS compared with CZE.

Amino Acid Sequence↗

PubMed1994

Inefficacy of low-dose continuous oral etoposide in non-small cell lung cancer.

Etoposide is more active in small cell lung cancer when given over 5 days than as a single injection. To examine this concept further, we designed this Phase II study in NSCLC using continuous low-dose oral etoposide. We enrolled 19 patients with measurable disease and the standard eligibility criteria. 16 had no prior chemotherapy. Etoposide was given at a dose of 50 mg by mouth daily. The median duration of therapy was 63 days (14-212 days). Toxicity was mild myelosuppression and GI symptoms. Therapy was discontinued because of progression of disease in 13 patients, toxicity (GI) in 3 patients; intercurrent disease, self-removal, and other reasons in 1 patient each. No complete or partial responses were seen (95% CI: 0-17.6%). The median survival after entry into the trial was 159 days (41-571+ days). We conclude that low-dose continuous oral etoposide is a well-tolerated but ineffective regimen in non-small cell lung cancer.

Administration, Oral↗