Surrogate System Research Articles

Assessment of the ultrastructural and proliferative properties of human embryonic stem cell-derived cardiomyocytes.

Assessment of early ultrastructural development and cell-cycle regulation in human cardiac tissue is significantly hampered by the lack of a suitable in vitro model. Here we describe the possible utilization of human embryonic stem cell (ES) lines for investigation of these processes. With the use of the embryoid body (EB) differentiation system, human ES cell-derived cardiomyocytes at different developmental stages were isolated and their histomorphometric, ultrastructural, and proliferative properties were characterized. Histomorphometric analysis revealed an increase in cell length, area, and length-to-width ratio in late-stage EBs (>35 days) compared with early (10-21 days) and intermediate (21-35 days) stages. This was coupled with a progressive ultrastructural development from an irregular myofibrillar distribution to an organized sarcomeric pattern. Cardiomyocyte proliferation, assessed by double labeling with cardiac-specific antibodies and either [3H]thymidine incorporation or Ki-67 immunolabeling, demonstrated a gradual withdrawal from cell cycle. Hence, the percentage of positively stained nuclei in early-stage cardiomyocytes ([3H]thymidine: 60 +/- 10%, Ki-67: 54 +/- 23%) decreased to 36 +/- 7% and 9 +/- 16% in intermediate-stage EBs and to <1% in late-stage cardiomyocytes. In conclusion, a reproducible temporal pattern of early cardiomyocyte proliferation, cell-cycle withdrawal, and ultrastructural maturation was noted in this model. Establishment of this unique in vitro surrogate system may allow to examine the molecular mechanisms underlying these processes and to assess interventions aiming to modify these properties. Moreover, the detailed characterization of the ES cell-derived cardiomyocyte may be crucial for the development of future cell replacement strategies aiming to regenerate functional myocardium.

Ectopic expression of a Haemonchus contortus GATA transcription factor in Caenorhabditis elegans reveals conserved function in spite of extensive sequence divergence.

Comparative analysis between Caenorhabditis elegans and other nematode species offers a powerful approach to study gene function. C. elegans also has great potential as a surrogate expression system to study the function of genes from parasitic nematode species where transgenic methodologies are unavailable. However there is little information on the extent to which the biology of C. elegans is conserved with other nematode species and very few parasitic nematode genes have yet been functionally expressed in C. elegans. We have identified and characterised a homologue of the C. elegans GATA transcription factor elt-2, a central regulator of endoderm development, from the parasitic nematode Haemonchus contortus. The H. contortus ELT-2 polypeptide is present in endoderm nuclei throughout embryonic and post-embryonic development, except for in the infective L3 stage, and our experiments reveal that the development of the H. contortus endodermal lineage is strikingly similar to that of C. elegans. Sequence conservation between the H. contortus and C. elegans ELT-2 polypeptides broadly reflects function since the major region of sequence identity corresponds to the DNA binding domain. However, the overall level of sequence identity is remarkably low with the only other major region of identity corresponding to an unusual zinc finger domain. In spite of this, ectopic expression of the H. contortus elt-2 gene in transgenic C. elegans is sufficient to activate a programme of endodermal differentiation demonstrating that function is highly conserved. This approach of ectopic expression using an inducible promoter provides an effective way in which to use C. elegans for the in vivo functional analysis of parasitic nematode genes.

Expression of Cpgp40/15 in Toxoplasma gondii: a surrogate system for the study of Cryptosporidium glycoprotein antigens.

Cryptosporidium parvum is a waterborne enteric coccidian that causes diarrheal disease in a wide range of hosts. Development of successful therapies is hampered by the inability to culture the parasite and the lack of a transfection system for genetic manipulation. The glycoprotein products of the Cpgp40/15 gene, gp40 and gp15, are involved in C. parvum sporozoite attachment to and invasion of host cells and, as such, may be good targets for anticryptosporidial therapies. However, the function of these antigens appears to be dependent on the presence of multiple O-linked alpha-N-acetylgalactosamine (alpha-GalNAc) determinants. A eukaryotic expression system that would produce proteins bearing glycosylation patterns similar to those found on the native C. parvum glycoproteins would greatly facilitate the molecular and functional characterization of these antigens. As a unique approach to this problem, the Cpgp40/15 gene was transiently expressed in Toxoplasma gondii, and the expressed recombinant glycoproteins were characterized. Antisera to gp40 and gp15 reacted with the surface membranes of tachyzoites expressing the Cpgp40/15 construct, and this reactivity colocalized with that of antiserum to the T. gondii surface protein SAG1. Surface membrane localization was dependent on the presence of the glycophosphatidylinositol anchor attachment site present in the gp15 coding sequence. The presence of terminal O-linked alpha-GalNAc determinants on the T. gondii recombinant gp40 was confirmed by reactivity with Helix pomatia lectin and the monoclonal antibody 4E9, which recognizes alpha-GalNAc residues, and digestion with alpha-N-acetylgalactosaminidase. In addition to appropriate localization and glycosylation, T. gondii apparently processes the gp40/15 precursor into the gp40 and gp15 component glycopolypeptides, albeit inefficiently. These results suggest that a surrogate system using T. gondii for the study of Cryptosporidium biology may be useful.

Enzymatic activities of the GB virus-B RNA-dependent RNA polymerase

The GB virus-B (GBV-B) nonstructural protein 5B (NS5B) encodes an RNA-dependent RNA polymerase (RdRp) with greater than 50% sequence similarity to the hepatitis C virus (HCV) NS5B. Recombinant GBV-B NS5B was reported to possess RdRp activity (W. Zhong et al., 2000, J. Viral Hepat. 7, 335–342). In this study, the GBV-B RdRp was examined more thoroughly for different RNA synthesis activities, including primer-extension, de novo initiation, template switch, terminal nucleotide addition, and template specificity. The results can be compared with previous characterizations of the HCV RdRp. The two RdRps share similarities in terms of metal ion and template preference, the abilities to add nontemplated nucleotides, perform both de novo initiation and extension from a primer, and switch templates. However, several differences in RNA synthesis between the GBV-B and HCV RdRps were observed, including (i) optimal temperatures for activity, (ii) ranges of Mn 2+ concentration tolerated for activity, and (iii) cation requirements for de novo RNA synthesis and terminal transferase activity. To assess whether the recombinant GBV-B RdRp may represent a relevant surrogate system for testing HCV antiviral agents, two compounds demonstrated to be active at nanomolar concentrations against HCV NS5B were tested on the GBV RdRp. A chain terminating nucleotide analog could prevent RNA synthesis, while a nonnucleoside HCV inhibitor was unable to affect RNA synthesis by the GBV RdRp.

Expression profiling of blood samples from an SU5416 Phase III metastatic colorectal cancer clinical trial: a novel strategy for biomarker identification.

BackgroundMicroarray-based gene expression profiling is a powerful approach for the identification of molecular biomarkers of disease, particularly in human cancers. Utility of this approach to measure responses to therapy is less well established, in part due to challenges in obtaining serial biopsies. Identification of suitable surrogate tissues will help minimize limitations imposed by those challenges. This study describes an approach used to identify gene expression changes that might serve as surrogate biomarkers of drug activity.MethodsExpression profiling using microarrays was applied to peripheral blood mononuclear cell (PBMC) samples obtained from patients with advanced colorectal cancer participating in a Phase III clinical trial. The PBMC samples were harvested pre-treatment and at the end of the first 6-week cycle from patients receiving standard of care chemotherapy or standard of care plus SU5416, a vascular endothelial growth factor (VEGF) receptor tyrosine kinase (RTK) inhibitor. Results from matched pairs of PBMC samples from 23 patients were queried for expression changes that consistently correlated with SU5416 administration.ResultsThirteen transcripts met this selection criterion; six were further tested by quantitative RT-PCR analysis of 62 additional samples from this trial and a second SU5416 Phase III trial of similar design. This method confirmed four of these transcripts (CD24, lactoferrin, lipocalin 2, and MMP-9) as potential biomarkers of drug treatment. Discriminant analysis showed that expression profiles of these 4 transcripts could be used to classify patients by treatment arm in a predictive fashion.ConclusionsThese results establish a foundation for the further exploration of peripheral blood cells as a surrogate system for biomarker analyses in clinical oncology studies.

Generalized Surrogate Problem Methodology for Online Stochastic Discrete Optimization

We consider stochastic discrete optimization problems where the decision variables are nonnegative integers and propose a generalized surrogate problem methodology that modifies and extends previous work in Ref. 1. Our approach is based on an online control scheme which transforms the problem into a surrogate continuous optimization problem and proceeds to solve the latter using standard gradient-based approaches while simultaneously updating both the actual and surrogate system states. In contrast to Ref. 1, the proposed methodology applies to arbitrary constraint sets. It is shown that, under certain conditions, the solution of the original problem is recovered from the optimal surrogate state. Applications of this approach include solutions to multicommodity resource allocation problems; in these problems, exploiting the convergence speed of the method, one can overcome the obstacle posed by the presence of local optima.

Online Surrogate Problem Methodology for Stochastic Discrete Resource Allocation Problems

We consider stochastic discrete optimization problems where the decision variables are nonnegative integers. We propose and analyze an online control scheme which transforms the problem into a surrogate continuous optimization problem and proceeds to solve the latter using standard gradient-based approaches, while simultaneously updating both the actual and surrogate system states. It is shown that the solution of the original problem is recovered as an element of the discrete state neighborhood of the optimal surrogate state. For the special case of separable cost functions, we show that this methodology becomes particularly efficient. Finally, convergence of the proposed algorithm is established under standard technical conditions; numerical results are included in the paper to illustrate the fast convergence of this approach.

Studies in Hebrew Narrative and Poetry

Studies in Hebrew Narrative and Poetry, by David Jobling. Berit Olam. Collegeville, MN: Liturgical Press, 1998. Pp. x + 330. $34.95. In this contribution to a fascinating commentary series, Jobling examines 1 Samuel according to T. Eagleton's triptych of class, race, and gender, exploring tension between 1 Samuel as a book in itself and as a part of a larger whole. Jobling's approach to 1 Samuel as Israel's national autobiography is a summation of course of his own academic career. The 1970s saw him engage structuralism and feminism, while poststructuralism and ideological criticism marked his 1980s work. In 1990s, he turned to new historicism and psychoanalysis. All of these methods come together in an analysis that affords Jobling personal reflection on his own career in a seminary with a high percentage of female students. Jobling denies that ideological criticism can be divorced from fight for equality for disempowered. He complains of travesty and narrowtology of new literary orthodoxy, naming specifically R. Alter, F. Kermode, M, Sternberg, L. Eslinger and especially R. Polzin (e.g., pp. 25, 144, 166-69, 308-9). Jobling replaces these scholars' concern for identifying literary artistry and narratorial omniscience in I Samuel with a program of uncovering points at which text reveals uncertainty over of past, and of recovering incompletely suppressed expressions of Israel's premonarchic egalitarian ideals. In ch. 2 Jobling divides 1 Samuel according to book divisions presumably original to DtrH (following M. Noth and D. J. McCarthy): Judg 2:11-1 Sam 12 is labeled Extended of Judges (hereafter EBJ) and I Sam 13-2 Sam 7 becomes Book of Everlasting Covenant (hereafter BEC). Jobling cogently argues that later synthesis of canonical book of 1 Samuel from DtrH has a great influence on how text is now interpreted, especially in terms of separating Hannah's story from those of women in Judges, and of changing emphasis in closing episodes. Class is discussed in part 2 (chs. 3-5) under guise of a critique of transition from judgeship to kingship in EBJ and BEC and then in canonical 1 Samuel. Jobling himself regards any form of kingship as a disaster and Israel's experience of it as a tragedy (p. 65). He asserts that 1 Samuel has no effective class analysis because it upholds one monarchic ideal as distinct from others, without realizing that all monarchies have specific social effects (p. 104). The canonical 1 Samuel develops idea of a surrogate fatherhood system of succession (Eli/Samuel, Samuel/Saul, and Saul /David) that avoids pitfalls of both judgeship and monarchy, although Jobling gives insufficient reasons why such a system was not also expressed by EBJ and BEC, both of which are components of a greater historical work. Part 3 (chs. 6-8) treats gender issues. Hannah is viewed according to a recuperative feminist paradigm. She is portrayed as a decisive character who makes connections between her unhappy life and national situation, especially regarding abuse of women at shrine. Attention is also drawn to cooption of Hannah by Deuteronomists, canonizers, and modem scholars. Jobling complains of -perverse royalist interpretations of Hannah's song that emphasize favorable mention of God's king in last verse (1 Sam 2:10) (p. 166). These interpretations reverse the meaning of song and trivialize it, character Hannah, and feminist biblical scholarship as a whole. Such interpreters also become complicit in Bible's destructive cultural effects (p. 169). This is certainly an unreasonable accusation, and, in any case, appeals to plain meaning of a text are not easily compatible with a deconstructive analysis. …

Development of a primary tamarin hepatocyte culture system for GB virus-B: a surrogate model for hepatitis C virus.

GB virus-B (GBV-B) causes an acute hepatitis in tamarins characterized by increased alanine transaminase levels that quickly return to normal as the virus is cleared. Phylogenetically, GBV-B is the closest relative to hepatitis C virus (HCV), and thus GBV-B infection of tamarins represents a powerful surrogate model system for the study of HCV. In this study, the course of infection of GBV-B in tamarins was followed using a real-time 5' exonuclease (TaqMan) reverse transcription-PCR assay to determine the level of GBV-B in the serum. Peak viremia levels exceeded 10(9) genome equivalents/ml, followed by viral clearance within 14 to 16 weeks. Rechallenge of animals that had cleared infection resulted in viremia that was limited to 1 week, suggestive of a strong protective immune response. A robust tissue culture system for GBV-B was developed using primary cultures of tamarin hepatocytes. Hepatocytes obtained from a GBV-B-infected animal maintained high levels of cell-associated viral RNA and virion secretion for 42 days of culture. In vitro infection of normal hepatocytes resulted in rapid amplification of cell-associated viral RNA and secretion of up to 10(7) genome equivalents/ml of culture supernatant. In addition, infection could be monitored by immunofluorescence staining for GBV-B nonstructural NS3 protein. This model system overcomes many of the current obstacles to HCV research, including low levels of viral replication, lack of a small primate animal model, and lack of a reproducible tissue culture system.

Adoptive transfer of genetically modified human hematopoietic stem cells into preimmune canine fetuses

To develop a surrogate model system for assaying gene transfer into human hematopoietic stem cells (HSCs) with in vivo repopulating potential, we injected human marrow cells transduced with a reporter retroviral vector in long-term marrow cultures (LTMCs), into the yolk sacs of preimmune canine fetuses. Of eight mid-gestation fetuses injected through the exteriorized uterine wall and under ultrasound guidance, seven were born alive. One puppy died in the neonatal period accidentally. The remaining six puppies are all healthy at 31 months of age. There was no evidence for graft-versus-host disease or any untoward effects of in utero adoptive transfer of transduced human LTMC cells. All puppies were chimeras. Human cells, detected by fluorescence in situ hybridization, were present in blood, declining from 38% to 0.05% between 10 and 44 weeks after birth. Corresponding numbers for marrow were from 20% to 0.05%. Human cells were also detected in assays of hematopoietic cell progenitors and in stimulated blood cultures. All six puppies were positive for the presence of proviral DNA at various time-points after birth. In three dogs, provirus was detected up to 41 weeks after birth in blood or marrow, and in one dog up to 49 weeks in blood. These data support the further development of this large-animal model system for studies of human hematopoie-sis.

Need money? roll the dice

In 1990, the Mississippi Legislature legalized dockside casino gaming as a surrogate tax system for the state. Since the establishment of the first casino in 1992, the casino industry has become an important component in the state's revenue picture. During fiscal years 1993, 1994, and 1995, gaming taxes and fees accounted for 1.6, 4.1, and 5.04 percent of Mississippi's general fund revenue, respectively. In addition, certain localities have benefitted from the industry as well. Beyond the direct revenue received through local taxation, casino gaming has substantially increased both property tax revenue and sales tax collections. The Mississippi Gaming Commission believes, however, that long-term revenue stability depends on developing resort destinations as opposed to a continued reliance on day-trippers. Colonists who balked at paying taxes to local governments were willing to purchase lottery tickets by those same governments(1) (yesterday). Government no longer possesses the capacity to secure the cooperation of the citizenry in meeting the requirements of public need. Consequently, it uses a buffer to take resources from the citizenry without invoking their wrath(2) (today).

A novel thrombopoietin signaling defect in polycythemia vera platelets.

The pathogenesis of polycythemia vera (PV), a disease involving a multipotent hematopoietic progenitor cell, is unknown. Thrombopoietin (TPO) is a newly characterized hematopoietic growth factor which regulates the production of multipotent hematopoietic progenitor cells as well as platelets. To evaluate the possibility that an abnormality in TPO-mediated signal transduction might be involved in the pathogenesis of PV, we examined TPO-induced protein tyrosine phosphorylation using platelets as a surrogate model system. Platelets were isolated from the blood of patients with PV as well as from patients with other chronic myeloproliferative disorders and control subjects. Impaired TPO-mediated platelet protein tyrosine phosphorylation was a consistent observation in patients with PV as well as those with idiopathic myelofibrosis (IMF), in contrast to patients with essential thrombocytosis, chronic myelogenous leukemia, secondary erythrocytosis, iron deficiency anemia, hemochromatosis, or normal volunteers. Thrombin-mediated platelet protein tyrosine phosphorylation was intact in PV platelets as was expression of the appropriate tyrosine kinases and their cognate substrates. However, expression of the platelet TPO receptor, Mpl, as determined by immunoblotting, chemical crosslinking or flow cytometry was markedly reduced or absent in 34 of 34 PV patients and also in 13 of 14 IMF patients. Impaired TPO-induced protein tyrosine phosphorylation in PV and IMF platelets was uniformly associated with markedly reduced or absent expression of Mpl. We conclude that reduced expression of Mpl is a phenotypic characteristic of platelets from patients with PV and IMF. The abnormality appears to distinguish PV from other forms of erythrocytosis and may be involved in the platelet function defect associated with PV.

Baculovirus merozoite surface protein 1 C-terminal recombinant antigens are highly protective in a natural primate model for human Plasmodium vivax malaria.

A successful anti-blood stage malaria vaccine trial based on a leading vaccine candidate, the major merozoite surface antigen-1 (MSP1), is reported here. The trial was based on Plasmodium cynomolgi, which is a primate malaria parasite which is highly analogous to the human parasite Plasmodium vivax, in its natural host, the toque monkey, Macaca sinica. Two recombinant baculovirus-expressed P. cynomolgi MSP1 proteins, which are analogous to the 42- and 19-kDa C-terminal fragments of P. falciparum MSP1, were tested by immunizing three groups of three animals each with either p42, p19, or both together. The vaccines were delivered subcutaneously in three doses at 4-week intervals with complete and incomplete Freund's adjuvants. Very high antibody titers were obtained against both vaccinating antigens as measured by enzyme-linked immunosorbent assay (10[6] and above) and against whole parasites as measured by indirect immunofluorescence assay (>10[5]), achieving, in most animals, about a 10-fold increase from the first to the last immunization. A blood stage challenge with P. cynomolgi parasites led, in three adjuvant-treated and three naive control animals, to blood infections which were patent for at least 44 days, reaching peak densities of 0.6 and 3.8%, respectively. In contrast, all except one of the nine animals in the three vaccinated groups were highly protected, showing either no parasitemia at all or transient parasitemias which were patent for only 1 or 2 days. When the three p19-vaccinated monkeys were rechallenged 6 months later, the protective efficacy was unchanged. The success of this trial, and striking analogies of this natural host-parasite system with human P. vivax malaria, suggests that it could serve as a surrogate system for the development of a human P. vivax malaria vaccine based on similar recombinant analogs of the P. vivax MSP1 antigen.

Influence of solvent effects on retention for a porous polymer sorbent in reversed phase liquid chromatography

The influence of acetonitrile, methanol and isopropanol as retention selectivily modifiers in reversed phase liquid chromatography on a poly(styrene-divinylbenzene) macroporous polymer sorbent (PLRP-S) is evaluated using the solvation parameter model. Retention results from a combination of adsorption and partitioning and is influenced by the equilibrium absorption of organic solvent by the polymer from the mobile phase. The sorption of solutes is dominated by the ease of cavity formation in or on the solvated sorbent, with a small contribution from lone pair-lone pair electron interactions. All polar interactions, such as dipole-type and hydrogenbond formation, are more favorable in the mobile phase and reduce retention. Changes in the uptake of organic solvent from the mobile phase affect kinetic properties of the column such as band broadening and porosity as well as retention. The PLRP-S solvated sorbent is suitable for solid-phase extraction and is more retentive than typical silica-based, bonded phase sorbents for extraction from water. As a surrogate system for estimating solute lipophilicity and biological activity through retention-property correlations it provides a poor fit for hydrogen-bond acid solutes and is too dipolar/polarizable to fit some models.

Durable power of attorney and informed consent with Alzheimer's disease patients: a clinical study.

Experience with a new surrogate consent system for patients with Alzheimer's disease is reviewed. It was hypothesized that as patients' cognitive status deteriorated, surrogate consent through a durable power of attorney would become necessary to facilitate continued involvement in clinical research. The authors retrospectively reviewed the charts of inpatients with Alzheimer's disease who participated in research between January 1989 and December 1994 at the Geriatric Psychiatry Unit of the National Institute of Mental Health. Seventy-nine patients were included. The main outcome measures were the Clinical Dementia Rating, Global Deterioration Scale for primary degenerative dementia, and Mini-Mental State. Most patients were in the mild-to-moderate stage of the illness when they chose to participate in research and assign a durable power of attorney (96% scored 2 or less on the Clinical Dementia Rating, and 92% scored 5 or less on the Global Deterioration Scale). On average, the subjects participated in 3.8 (SD = 2.6) studies. For 35 patients with multiple admissions over this period (average = 3.1 years), scores on the Clinical Dementia Rating and Global Deterioration Scale declined by 1.0 and 1.5 points, respectively. The durable power of attorney allows research participation for subjects with Alzheimer's disease at all stages. The linchpin is assignment of a durable power of attorney in the early-to-moderate stage of Alzheimer's disease, before subjects lose the capacity to give informed consent. This approach could also be adapted to patients with cognitive decline due to other debilitating diseases.

A generalized approach to the control of the evolution of a molecular system

The theory of active control of molecular motion by use of shaped laser pulses is developed emphasizing the role of interference and using thermodynamic analogies. Attention is focused on the control of the dynamics in a system with n states coupled by radiation, and the phase relations which generate particular control schemes are derived. Among the new results reported is an optimal control scheme which constrains the value of the phase. The n-state model can be considered to represent a molecule with n electronic potential energy surfaces and an arbitrary number of degrees of freedom or as the skeleton spectrum of system where each level in the spectrum can be associated with a specific set of quantum numbers for all of the degrees of freedom. We show how the control of the dynamics of an n-state molecule can be represented in terms of the control of the dynamics of a precisely defined surrogate fewer state system. This reduction is illustrated by use of a surrogate two state system to describe the dynamics of population transfer in a three state system.

Accessibility of the fluorescent reporter group in native, silica-adsorbed, and covalently attached acrylodan-labeled serum albumins.

Fluorescence quenching techniques are used to investigate the accessibility of a model biorecognition element-reporter group system when in buffer, surface-adsorbed, and covalently attached to a silica surface. The site-selective fluorescent reporter group, 6-acryloyl(dimethylamino)naphthalene (acrylodan, Ac), is attached covalently (at cysteine-34) to bovine and human serum albumin (BSA and HSA, respectively) and serves as a surrogate recognition element-reporter group system. Molecular oxygen is used to quench the Ac fluorescence and the accessibility, in the form of bimolecular rate constants (kq), in each model system is quantified. Although one might expect these systems to exhibit similar behavior, differences in quenching characteristics are observed, such as wavelength dependency of the Stern-Volmer quenching constant (KSV) for the native proteins in buffer. BSA-Ac exhibits wavelength dependent KSV values as well as a blue-shifted emission spectrum on O2 addition. Physisorption of BSA-Ac onto a fused-silica optical fiber lowers the accessibility of Ac to O2, whereas covalent attachment of BSA-Ac to APTES/glutaraldehyde-modified silica enhances the accessibility of the Ac reporter group to O2.

Acetylcholinesterase expression in NTera 2 human neuronal cells: a model for developmental expression in the nervous system.

Acetylcholinesterase (AChE; EC 3.1.1.7) is expressed in the central nervous system in multiple molecular forms that may subserve multiple functions and may be selectively lost in neurodegenerative illnesses such as Alzheimer's disease. AChE expression has been studied in primary cultures of developing vertebrate nervous system, but investigation has been limited by the lack of a suitable human CNS surrogate cell model system for in vitro studies and the inability of primary brain cultures to provide large numbers of pure neurons. To develop an in vitro model for studies of neuronal AChE expression and function, we utilized a neuronally committed human teratocarcinoma cell line, NTera 2, that can be induced to differentiate to a post-mitotic CNS neuronal phenotype. We found that NTera 2 cells express multiple molecular forms of AChE that are similar to CNS-derived AChE isoforms in velocity sedimentation profile, anion exchange elution profile, and sensitivity to inhibitors. At least two forms of AChE are expressed (G1 and G4), similar to human and rodent brain, and induction of NTera 2 cell differentiation results in an increased G4/G1 ratio, which is characteristic of mature neurons. As in primary CNS neurons, AChE is present in NTera 2 cells in both the cytosolic fraction and in the outer membrane, and is also released in a soluble form. These observations indicate that NTera 2 cells provide a useful human model system for studies of expression of cell-associated and soluble cell-free AChE in developing and mature human neurons and for elucidating the potential role(s) of acetylcholinesterase metabolism in both normal development and neurodegenerative disease states.

Feasibility of Organo-Beryllium Target Mandrels Using Organo-Germanium PECVD as a Surrogate

Inertial Confinement Fusion capsules incorporating beryllium are becoming attractive for use in implosion experiments designed for modest energy gain. This paper explores the feasibility of chemical vapor deposition of organo-beryllium precursors to form coating materials of interest as ablators and fuel containers. Experiments were performed in a surrogate chemical system utilizing tetramethylgermane as the organometallic precursor. Coatings with up to 60 mole percent germanium were obtained. These coatings compare favorably with those previously reported in the literature and provide increasing confidence that a similar deposition process with an organo-beryllium precursor would be successful.

Approximation for the variance of the waiting time in a GI/G/1 queue

Obtaining a computable expression for the variance of the waiting time in a GI/G/1 queue is extremely difficult. We use the concept of isomorphism in this paper to develop such an approximation. The concept of isomorphism consists of identifying a surrogate system which has some points of similarity with a given system. Such an idea may be used in the applications involving reliability, inventory, queueing and insurance systems. In this paper, we replace a GI/G/1 system with an equivalent M/G/1 system and use the expression for the variance of the waiting time in an M/G/1 system to approximate the variance in a GI/G/1 system. We present simulation results to demonstrate the quality of our approximation.