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Biomedical subjects

A Lu

Publications and source records attributed to A Lu.

At least 73 records · Page 4Linked to original sources
PubMed1995

Immune responses induced by administration of encapsidated poliovirus replicons which express HIV-1 gag and envelope proteins.

Several viruses have been exploited for the development of recombinant vaccine vectors in which to express foreign proteins. Recently, we have described a system utilizing the RNA virus, poliovirus. We have constructed poliovirus genomes in which regions of the capsid have been substituted with gene fragments of the HIV gag and env genes. A complementation system has been designed to encapsidate defective genomes by providing the capsid protein in trans from a recombinant vaccinia virus (VV-P1). Serial passage in the presence of VV-P1 resulted in the generation of stocks of these encapsidated replicons. Infection of cells with these encapsidated replicons resulted in the expression of the recombinant protein as a fusion protein with the poliovirus capsid proteins VP4 and VP1. In this study, we have utilized encapsidated replicons which express the HIV-1-gag capsid protein (p24) as well as 1.5 kb of the HIV-1 env gene. Stocks of these encapsidated replicons were obtained by 20 serial passages in the presence of VV-P1. In addition, passage of the encapsidated replicons in the presence of poliovirus type 2 Lansing resulted in the encapsidation of the replicons by the capsid proteins provided by poliovirus. The administration of the type 2 Lansing/encapsidated replicons expressing HIV-1 gag in BALB/c mice by intramuscular, intrarectal, or intragastric routes resulted in the generation of antibodies in the serum and secretions against both poliovirus and HIV-1 gag. To prove that the replicons alone are immunogenic, we administered replicons expressing either HIV-1 gag or env to transgenic mice which expressed the receptor for poliovirus type 1. Immunization of these mice by the intramuscular route resulted in the generation of serum antibodies specific for poliovirus as well as for HIV-1 antigens. The results obtained led us to the conclusion that the replicons are immunogenic when given alone or in the presence of poliovirus. These results are important for the use of the poliovirus replicons as a recombinant vaccine vector.

AIDS Vaccines↗
PubMed1995

Pulmonary function testing in spinal cord injury: correlation with vital capacity.

Spinal cord injury (SCI) causes restrictive ventilatory changes, with reductions in vital capacity, functional residual capacity, and expiratory reserve volume. Vital capacity (VC) often is used as an indicator of overall pulmonary function in these patients. In an effort to determine the extent to which VC correlates with other pulmonary function tests, 52 patients with recent acute traumatic SCI underwent complete pulmonary function testing. Statistical relationships were determined between VC and nine other tests. VC was found to be significantly correlated with forced expiratory volume in 1 s, inspiratory capacity, expiratory reserve volume, functional residual capacity, residual volume (RV), total lung capacity (TLC), and RV/TLC ratio, but not with maximum positive expiratory pressure nor with maximum negative inspiratory pressure. The excellent correlations between vital capacity and nearly all of the other pulmonary function tests support the use of VC as a single global measure of overall ventilatory status in SCI patients.

Adolescent↗
PubMed1995

Differential requirements for baculovirus late expression factor genes in two cell lines.

A plasmid library of 18 late expression factor (LEF) genes (LEF library) from the baculovirus Autographa californica nuclear polyhedrosis virus (AcMNPV) supports transient expression from a late viral promoter in the SF-21 cell line, derived from Spodoptera frugiperda. We found, however, that this LEF library was unable to support expression from the same promoter in the TN-368 cell line, derived from Trichoplusia ni, which is also permissive for AcMNPV replication. To identify the additional factor(s) required for expression in TN-368 cells, we cotransfected the LEF library with clones representing portions of the AcMNPV genome not represented in the LEF library. A single additional gene was identified; this gene corresponded to ORF70 of the complete AcMNPV sequence and potentially encodes a 34-kDa cysteine-rich polypeptide. Because of its differential effect on late gene expression in the two cell lines, we renamed ORF70 hcf-1 (for host cell-specific factor 1). hcf-1 was involved in expression from reporter plasmids under late and very late but not early promoter control, indicating that it was also a LEF gene. Plasmid DNA replication assays indicated that HCF-1 was involved in virus origin-specific DNA replication in TN-368 cells. Three LEF genes, ie-2, lef-7, and p35, required for optimal virus origin-specific plasmid DNA replication or stability in SF-21 cells had little or no influence in TN-368 cells. Thus, as determined by transient-expression assays, cell line-specific and potentially host-specific factors are required for origin-specific DNA replication or stability.

Animals↗
PubMed1995

The roles of eighteen baculovirus late expression factor genes in transcription and DNA replication.

A set of 18 plasmid subclones of the Autographa californica nuclear polyhedrosis virus genome supports expression from a late viral promoter in transient expression assays (J. W. Todd, A. L. Passarelli, and L. K. Miller, J. Virol. 69:968-974, 1995). Using this set of plasmids, we have assigned a role for each of the 18 genes required for optimal late gene expression with respect to its involvement at the levels of transcription, translation, and/or DNA replication. RNase protection analyses demonstrated that all of the known late expression factor genes (lefs) affected the steady-state level of reporter gene RNA. Thus, none of the lefs appeared to be specifically involved in translation. A subset of the lefs supported plasmid replication; ie-1, lef-1, lef-2, lef-3, p143, and p35 were essential for plasmid replication, while ie-n, lef-7, and dnapol had stimulatory effects. The predicted sequence of lef-7 suggests that it is a homolog of herpesvirus single-stranded DNA-binding protein (UL29). The role of p35 in plasmid replication appears to be suppression of apoptosis, because p35 could be functionally replaced in the replication assay by either Cp-iap or Op-iap, two heterologous baculovirus genes which suppress apoptosis by a mechanism which appears to differ from that of p35. Thus, one or more of the replication-related lefs or the process of plasmid replication appears to induce cellular apoptosis. Our results indicate that the remaining lefs, lefs 4 through 11, p47, and 39K (pp31), function either at the level of transcription or at that of mRNA stabilization.

Amino Acid Sequence↗
PubMed1994

A rapid and efficient one-tube PCR-based mutagenesis technique using Pfu DNA polymerase.

A rapid method for efficiently generating site-directed mutations on a clean sequence background is described. This modification of the megaprimer PCR mutagenesis approach can be performed in one tube in less than 4.5 hours, and does not require purification of intermediate products. High fidelity of DNA sequence replication is obtained by employing Pfu DNA polymerase and limiting the total number of amplification cycles to 30. The mutagenesis efficiency of the procedure is high enough to allow rapid, direct identification of mutants by restriction digest or sequencing techniques.

DNA-Directed DNA Polymerase↗
PubMed1994

Identification of three late expression factor genes within the 33.8- to 43.4-map-unit region of Autographa californica nuclear polyhedrosis virus.

A transient transactivation assay system was used in combination with an overlapping Autographa californica nuclear polyhedrosis virus clone library to identify genes involved in late and very late baculovirus gene expression. We have identified three genes within the 33.8- to 43.4-map-unit region of the A. californica nuclear polyhedrosis virus genome which contribute to expression from promoters of the vp39 major capsid protein and polyhedrin genes. One of these three genes corresponds to the previously identified DNA polymerase gene, while the other two genes encode previously unidentified polypeptides of 59,418 and 8,706 Da. None of these genes were required for expression from the early etl promoter.

Animals↗
PubMed1994

Getting the most out of low-dose syringe prefilling of a high-cost biotechnology agent.

The recent increased usage of high-cost biotechnology agents has placed a tremendous impact on the hospital pharmacy budget. One approach in improving cost containment is to minimize waste during the preparation of these agents. This is particularly practical and possible in the process of low-dose syringe repackaging of Sargramostim (GM-CSF) used for the treatment of neutropenia. In search of ways to reduce waste, this study looked into the dos and don'ts of repackaging this agent in syringes. Decreased waste is very significant if GM-CSF is properly reconstituted with bacteriostatic water for injection and if a syringe equipped with a permanently attached needle is used. Another 10% of the solution from each vial could be saved if the solution is withdrawn with a technique involving the vial in a right-side-up position.

Biotechnology↗
PubMed1994

[The experimental study of prevention and treatment of the side-effects of chemotherapy with acupuncture (comparison among the effect of acupuncture at different acupoint)].

After an intraperitoneal injection of cyclophosphamide in rats, there was pathologic changes in hemopoietic, immune and visceral function. Acupuncture respectively at Zusanli, Dazhui and Shenshu points could decrease the above mentioned harms in the rats, the effect of acupuncture on hemopoietic function: Dazhui more than or equal to Zusanli, Zusanli more than Shenshu; the effect of acupuncture on immune function: Zusanli more than Dazhui, Dazhui more than or equal to Shenshu; the effect of acupuncture on the function of the liver and the kidney: Shenshu more than or equal to Zusanli, Zusanli more than Dazhui. The effect of acupuncture at all three points, i.e. Zusanli. Dazhui and Shenshu was more than that at any single point due to their coordination. There was no obvious effect when acupuncture at non-acupoints on tail.

Acupuncture Points↗
PubMed1993

Immediate-early baculovirus genes transactivate the p143 gene promoter of Autographa californica nuclear polyhedrosis virus.

Trans-acting regulatory components of Autographa californica nuclear polyhedrosis virus (AcMNPV) were studied in a transient assay system for their ability to activate the p143 gene promoter. A region including 502 nt upstream of the AUG of the p143 gene was linked to the bacterial chloramphenicol acetyltransferase (CAT) gene. The p143 promoter-CAT construct was used to identify AcMNPV immediate-early gene products required for expression from the p143 gene early promoter. Transient expression assays in uninfected Spodoptera frugiperda cells indicated that the IE-1 gene product was capable of transactivating the p143 gene promoter and this activation was augmented by the IE-N gene product. In addition, another immediate-early gene, PE-38, was shown to mediate transactivation of the p143 gene promoter but not the 39K delayed early gene promoter. Deletions to -187 bp relative to the p143 RNA start site did not significantly affect promoter activity in the combined presence of IE-1 and the HindIII-F fragment. However, a deletion to -52 bp decreased promoter activity by 40%. In contrast, in the presence of ts8 DNA at 33 degrees, deletions to -52 bp decreased promoter activity by 80% suggesting that additional viral factors were involved in p143 gene promoter regulation.

Animals↗
PubMed1993

Effects of pentoxifylline on sperm motility and hyperactivation in normozoospermic and normokinetic semen.

OBJECTIVE: To determine the effects of in vitro incubation with pentoxifylline on sperm motion characteristics of spermatozoa from normozoospermic, normokinetic specimens. DESIGN: Prospective, controlled experiment. SETTING: Andrology laboratory, university-based fertility center. PARTICIPANTS: Healthy, untreated male partners of couples attending the fertility center. INTERVENTION: Each specimen was washed, pelleted by centrifugation, then resuspended in human tubal fluid medium (HTF). Two portions were incubated at 37 degrees C, one with pentoxifylline (final concentration = 1 mg/mL = 3.6 mM) and the other without pentoxifylline (control). After 1 hour, the pentoxifylline-treated portion was divided: one half was washed to remove pentoxifylline, then further incubated in HTF; the other half remained incubated in HTF with pentoxifylline. MAIN OUTCOME MEASURES: Motility, hyperactivation, amplitude of lateral head displacement (ALH), curvilinear velocity (VCL), straight line velocity (VSL), linearity (LIN), beat-cross frequency. RESULTS: Incubation with pentoxifylline did not increase motility, VSL, LIN, or beat-cross frequency but did significantly increase HA, VCL, and ALH at 1, 2, and 4 hours, compared with control. Incubation in control medium without pentoxifylline did not significantly increase any of the parameters measured. After 24 hours of incubation with or without pentoxifylline, all parameters measured were significantly decreased, with the exception of LIN. CONCLUSIONS: Pentoxifylline does not increase percentage motility of washed spermatozoa in capacitation medium. Enhancement of sperm HA by pentoxifylline in capacitation medium occurs with normozoospermic, normokinetic semen specimens. This effect persists for up to 4 hours when pentoxifylline is removed from the medium after 1 hour of incubation.

Analysis of Variance↗
PubMed1992

Nucleotide sequence and transcriptional analysis of the p80 gene of Autographa californica nuclear polyhedrosis virus: a homologue of the Orgyia pseudotsugata nuclear polyhedrosis virus capsid-associated gene.

The 67.2- to 68.5-m.u. region of Autographa californica nuclear polyhedrosis virus was sequenced. A large open reading frame (ORF) was identified in the clockwise direction on the circular genome map which could potentially encode an 80-kDa polypeptide. Analysis of the predicted amino acid sequence of this ORF indicated that it was a homologue of the p87 capsid-associated gene of Orgyia pseudotsugata MNPV with an overall amino acid similarity of 34.3%. A late transcript of 2.1 kb was mapped to this open reading frame. An antisense 3.1-kb transcript partially overlapped the 5' end of the 2.1-kb RNA. Anti-extracellular virus sera reacted with a fusion protein consisting of a portion of the AcMNPV p80 gene fused to the bacterial trpE gene product, suggesting that the AcMNPV p80 gene product was also a component of the virus capsid.

Amino Acid Sequence↗
PubMed1992

Silver-induced frog skeletal muscle contraction and its modulation by calcium antagonists nifedipine and felodipine and calcium agonist Bay K 8644.

1. Ag+ induces one phase of a transient contracture in frog skeletal muscle (0.3-10 microM) and potentiates twitch tension when the fiber is given continuous stimulation. 2. The potentiation of fiber contraction by Ag+ is similar to the effect of Ca2+ antagonists nifedipine and felodipine. 3. Bay K 8644 (100 nM) potentiates and accelerates Ag(+)-induced tension development and the inactivation occurs more rapidly than in the control (Ag+ alone). 4. Two factors can be considered to be essential for the induction of Ag+ contracture: (1) a certain number of Ag+ ions must bind to free SH groups of the voltage sensor; and (2) the binding must occur within a limited time to raise the mechanical threshold to induce contracture. 5. All results suggest that Ag+ binding to crucial SH groups on the Ca2+ channel may be responsible for the activation of muscle contraction, potentiation, and the inhibition of excitation-contraction coupling in skeletal muscle.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1992

The inhibitory effects of 21 mimics of superoxide dismutase on luminol-mediated chemiluminescence emitted from PMA-stimulated polymorphonuclear leukocyte.

Four groups comprising 21 superoxide dismutase (SOD) mimics synthesized by us were comparatively studied for their inhibitory effects on luminol-mediated chemiluminescence emitted from phorbol myristate acetate (PMA)-stimulated polymorphonuclear leukocyte (PMNL). Among these groups, 20-membered macrocyclic bicopper(II) complexes and 13-membered macrocyclic dioxotetramine copper(II) complexes exhibited relatively higher activities of scavenging reactive oxygen species (ROS) produced by PMA-stimulated PMNL as compared with polyamine Cu(II)-Zn(II) complexes and copper(II) complexes of bis-shiff-base. Moreover, distinctly different effects of SOD mimics in the biological system have been found even in the same group. It is suggested that the biological effects of some SOD mimics are related to their structures.

Chelating Agents↗
PubMed1992

Transcription analysis of the EcoRI D region of the baculovirus Autographa californica nuclear polyhedrosis virus identifies an early 4-kilobase RNA encoding the essential p143 gene.

We have investigated the transcriptional activity of the 60.1- to 68.3-map-unit region of the baculovirus Autographa californica nuclear polyhedrosis virus (AcMNPV). Twelve transcripts mapping to this region were expressed at various times during infection. An early 4.0-kb transcript, potentially coding for a 143-kDa peptide essential for viral DNA replication, was maximally abundant at 6 h postinfection (p.i.). Transcripts of 0.5, 1.1, 1.4, 2.1, and 3.1 kb were most abundant at 12 h p.i., while two large transcripts of 5.2 and 6.8 kb were expressed maximally at 24 h p.i. In the presence of cycloheximide, and in ts8-infected cells at the nonpermissive temperature, only the 4.0-kb RNA was expressed. Northern (RNA) blot analysis using DNA subfragments from the EcoRI D fragment as probes suggested that many of the transcripts overlapped. Strand-specific cRNA probes revealed that the majority of the RNAs were transcribed in the counterclockwise direction. S1 nuclease and primer extension analysis were used to map the 5' ends of transcripts coded within the 60.1- to 64.8-map-unit region. Mapping of the 3' ends of the 1.1-, 4.0-, 5.2-, and 6.8-kb transcripts suggested that these RNAs were all coterminal at their 3' ends. A minicistron was found between the early 4.0-kb transcription start site and the predicted ATG start codon of the p143 gene. Several similar sequence motifs were identified in the promoter regions of the p143 gene and the AcMNPV DNA polymerase gene.

Animals↗