Simulated microgravity-induced vascular hyporesponsiveness: role of signal transduction.
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Animals↗
Biomedical subjects
H Nguyen
Publications and source records attributed to H Nguyen.
Varicella-zoster virus ORF61 deletion mutants replicate in cell culture, but a mutant with stop codons in ORF61 reverts to wild-type virus.
Varicella-zoster virus (VZV) ORF61 encodes a phosphoprotein that transactivates VZV promoters. Transfection of cells with cosmid DNAs, including a cosmid with a large deletion in ORF61, resulted in a VZV ORF61 deletion mutant that was impaired for growth in vitro and could be partially complemented by growth in neuroblastoma or osteosarcoma cell lines. Cells infected with the VZV ORF61 deletion mutant expressed normal levels of an immediate-early VZV protein, but had reduced levels of a late protein and showed abnormal syncytia. Carboxy terminal truncation mutants of VZV ORF61 protein have a transrepressing phenotype and inhibit the infectivity of cotransfected wild-type viral DNA. Transfection of cells with cosmid DNAs, including a cosmid with stop codons that should result in an ORF61 truncation mutant expressing a transrepressing protein that retains the RING finger domain, resulted in a viral genome which reverted back to the wild-type sequence. BAL-31 exonuclease was used to produce deletions at the site of the stop codons in ORF61 of the cosmid, resulting in loss of the RING finger domain. Transfection of tissue culture cells with the ORF61 BAL-31 deletion mutants and other cosmid DNAs yielded viable viruses. Thus, while deletion mutants lacking the RING finger domain of ORF61 replicate in cell culture, a mutant with stop codons that retains this domain could not be propagated and reverted to wild-type virus.
Codon, Terminator↗
Hormone-dependent coactivator binding to a hydrophobic cleft on nuclear receptors.
The ligand-binding domain of nuclear receptors contains a transcriptional activation function (AF-2) that mediates hormone-dependent binding of coactivator proteins. Scanning surface mutagenesis on the human thyroid hormone receptor was performed to define the site that binds the coactivators, glucocorticoid receptor-interacting protein 1 (GRIP1) and steroid receptor coactivator 1 (SRC-1). The residues involved encircle a small surface that contains a hydrophobic cleft. Ligand activation of transcription involves formation of this surface by folding the carboxyl-terminal alpha helix against a scaffold of three other helices. These features may represent general ones for nuclear receptors.
HeLa Cells↗
Montelukast for chronic asthma in 6- to 14-year-old children: a randomized, double-blind trial. Pediatric Montelukast Study Group.
CONTEXT: Leukotrienes are important mediators of asthma by causing bronchoconstriction, mucous secretion, and increased vascular permeability. Studies using compounds that block leukotrienes have demonstrated improvement in asthma control in adults and adolescents, but children younger than 12 years, for whom asthma is the most common chronic disease, have not been studied. OBJECTIVE: To determine the clinical effect of montelukast, a leukotriene receptor antagonist, in 6- to 14-year-old children with asthma. DESIGN: Eight-week, multicenter, randomized, double-blind study. SETTING: Forty-seven outpatient centers at private practices and academic medical centers in the United States and Canada. PATIENTS: A total of 336 children with forced expiratory volume in 1 second (FEV1) between 50% to 85% of the predicted value, at least 15% reversibility after inhaled beta-agonist administration, a minimal predefined level of daytime asthma symptoms, and daily beta-agonist use. Concomitant inhaled corticosteroids at a constant daily dose were used by 39% of patients receiving montelukast and 33% receiving placebo. INTERVENTION: After a 2-week placebo run-in period, patients received either montelukast (5-mg chewable tablet) or matching-image placebo once daily at bed-time for 8 weeks. MAIN OUTCOME MEASURE: Morning FEV1 percent change from baseline. RESULTS: Mean morning FEV1 increased from 1.85 L to 2.01 L in the montelukast group and from 1.85 L to 1.93 L in the placebo group. This represents an 8.23% (95% confidence interval [CI], 6.33% to 10.13%) increase from baseline in the montelukast group and a 3.58% (95% CI, 1.29% to 5.87%) increase from baseline in the placebo group (P<.001 for montelukast vs placebo). CONCLUSION: Montelukast improves morning FEV1 in 6- to 14-year-old children with chronic asthma.
Acetates↗
A transgenic myogenic cell line lacking ryanodine receptor protein for homologous expression studies: reconstitution of Ry1R protein and function.
CCS embryonic stem (ES) cells possessing two mutant alleles (ry1r-/ry1r-) for the skeletal muscle ryanodine receptor (RyR) have been produced and injected subcutaneously into severely compromised immunodeficient mice to produce teratocarcinomas in which Ry1R expression is absent. Several primary fibroblast cell lines were isolated and subcloned from one of these tumors that contain the knockout mutation in both alleles and exhibit a doubling time of 18-24 h, are not contact growth inhibited, do not exhibit drastic morphological change upon serum reduction, and possess the normal complement of chromosomes. Four of these fibroblast clones were infected with a retrovirus containing the cDNA encoding myoD and a puromycin selection marker. Several (1-2 microg/ml) puromycin-resistant subclones from each initial cell line were expanded and examined for their ability to express myoD and to form multinucleated myotubes that express desmin and myosin upon removal of mitogens. One of these clones (1B5 cells) was selected on this basis for further study. These cells, upon withdrawal of mitogens for 5-7 d, were shown by Western blot analysis to express key triadic proteins, including skeletal triadin, calsequestrin, FK506-binding protein, 12 kD, sarco(endo)plasmic reticulum calcium-ATPase1, and dihydropyridine receptors. Neither RyR isoform protein, Ry1R (skeletal), Ry2R (cardiac), nor Ry3R (brain), were detected in differentiated 1B5 cells. Measurements of intracellular Ca2+ by ratio fluorescence imaging of fura-2-loaded cells revealed that differentiated 1B5 cells exhibited no responses to K+ (40 mM) depolarization, ryanodine (50-500 microM), or caffeine (20-100 mM). Transient transfection of the 1B5 cells with the full-length rabbit Ry1R cDNA restored the expected responses to K+ depolarization, caffeine, and ryanodine. Depolarization-induced Ca2+ release was independent of extracellular Ca2+, consistent with skeletal-type excitation-contraction coupling. Wild-type Ry1R expressed in 1B5 cells were reconstituted into bilayer lipid membranes and found to be indistinguishable from channels reconstituted from rabbit sarcoplasmic reticulum with respect to unitary conductance, open dwell times, and responses to ryanodine and ruthenium red. The 1B5 cell line provides a powerful and easily managed homologous expression system in which to study how Ry1R structure relates to function.
Animals↗
In vivo knee joint loading and kinematics before and after ACL transection in an animal model.
Osteoarthritis (OA) is typically diagnosed in humans in its final stage when joint movement becomes painful. Clinical information about the onset and the mechanisms triggering the degenerative responses are virtually non-existent. However, research on animal models of experimental OA shows that joint adaptations associated with the onset of OA can be detected as early as two to four weeks following disruption of the normal joint mechanics. Transection of the anterior cruciate ligament (ACL) has been shown to cause OA-like symptoms in various animal models including the cat. However. the changes in joint loading responsible for the early tissue responses have not been quantified in vivo. Consequently, the relationship between abnormal joint loading and the onset of OA remains unknown. The purpose of this study was to quantify knee loading before and early after ACL transection in the cat. Knee mechanics were assessed by measuring patellar tendon forces, gastrocnemius forces, knee flexor and extensor EMGs, and hindlimb kinematics before and 5, 7, and 9 days following ACL transection in six experimental and two sham-operated animals. The knee mechanics were not affected by sham-surgery but the muscular forces. knee extensor EMGs, and knee range of motion were reduced following ACL transection compared to corresponding pre-intervention values. These results suggest that ACL transection causes a general unloading and changed kinematics of the knee. We speculate that the decrease in loading and the altered kinematics are responsible for the onset of biologic adaptations of the knee. Precise data about the local joint contact mechanics before and after ACL transection are now required to further relate the detailed changes in the knee mechanics to the early joint changes.
Animals↗
One-year changes in hind limb kinematics, ground reaction forces and knee stability in an experimental model of osteoarthritis.
Long-term changes in the three-dimensional external loading, hind limb kinematics and knee stability were assessed in an anterior cruciate ligament (ACL)-transected cat model of osteoarthritis (OA). Seven skeletally mature cats (mean mass 4.6+/-1.4 kg) were studied before ACL transection (ACLT) and at 1 and 3 weeks, and at 3, 6, 9 and 12 months following ACLT. One week following ACLT, significant changes from the normal locomotion pattern were observed: peak vertical and anterior posterior ground reaction forces were decreased, particularly the peak posterior forces in the early phase of stance. Furthermore, knee angles were reduced by about 15 degrees throughout the whole gait cycle, while ankle and hip angles were reduced at paw off in the experimental compared to the contralateral hind limbs. Ground reaction forces and hind limb kinematics recovered to near pre-surgical patterns over the one year period assessed. ACLT was also associated with an increased knee instability which improved over time. X-rays suggested that there was a continued degeneration in the experimental knee over the one year period; there was osteophyte formation at the joint margins and an increase in cartilage thickness throughout the joint. It was speculated that the more flexed knee angles and the reduced anterior-posterior ground reaction forces in the ACL-transected compared to the intact hind limb represent an adaptive strategy aimed at avoiding excessive anterior displacement of the tibia in the early phase of stance. The recovery of the locomotion pattern with time might be related to the corresponding improvement of knee stability.
Animals↗
Microscopic evidence against HIV-1 infection of germ cells or attachment to sperm.
For a number of years we have intensively investigated the localization of HIV-1 in male genital tract tissues and secretions using a variety of microscopy techniques including immunocytochemistry, in situ hybridization, in situ PCR and electron microscopy. Our studies have failed to demonstrate an association between HIV-1 and either testicular germ cells or spermatozoa. In this article we present our results in the context of other related studies, and discuss the strengths and weaknesses of the techniques that have been used to address this important research question.
Animals↗
Administrative update: utilization of services. II. Utilization of mental health services in a tri-ethnic sample of adolescents.
In our study of a tri-ethnic sample of 2528 junior and high school students, we examined utilization of outpatient mental health services in relation to a number of variables cited in the literature as leading to potential biases and barriers to care. These include: age, gender, ethnicity, socioeconomic status, family size and composition, and linguistic fluency in Hispanic youth. The impact of service availability was examined through differences between the two regions studied: a well-served region of coastal southeast Texas and the markedly under served lower Rio Grande Valley. The impact of symptomatology was evaluated using the total problem score on the Youth Self Report by Achenbach. Hispanic youth had significantly lower mean service utilization than non-Hispanic whites. Multiple regression analyses demonstrated that socioeconomic status and family composition had a greater relative impact on utilization than all other non-clinical factors, both for the total sample as well as for the Hispanic sample. Ethnicity may play a significant role in child mental health services utilization through its close association to socioeconomic status.
Adolescent↗
Offenders with mental illness: prevalence and responsibility.
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Crime↗
Age-related changes in plasma and tissue fatty acid composition in Fischer 344 rats.
Advancing age is associated with increased risk of coronary artery disease. Changes in fatty acid metabolism affect important cellular membrane properties and functions which may contribute to the vascular pathophysiology of aging. This study was designed to investigate the effects of aging on the fatty acid composition of the plasma, liver, aorta, and renal artery in 4-, 15-, and 24-month old Fischer 344 rats, an animal model for aging. With aging, the levels of total polyunsaturated fatty acids (PUFA) increased in the plasma, aorta, and renal artery. The major changes in the liver fatty acid profile were increases in the levels of 18:2n6 and 18:3n3 and a decrease in the levels of 20:3n6 and 20:5n3. The results indicate that significant shifts occur in the levels of n6 and n3 PUFA in the plasma, liver, and vasculature with aging. The alterations in the fatty acid composition may be a pathogenetic mechanism of the vascular changes associated with aging.
Aging↗
Familial eosinophilia maps to the cytokine gene cluster on human chromosomal region 5q31-q33.
Familial eosinophilia (FE) is an autosomal dominant disorder characterized by peripheral hypereosinophilia of unidentifiable cause with or without other organ involvement. To localize the gene for FE, we performed a genomewide search in a large U.S. kindred, using 312 different polymorphic markers. Seventeen affected subjects, 28 unaffected bloodline relatives, and 8 spouses were genotyped. The initial linkage results from the genome scan provided evidence for linkage on chromosome 5q31-q33. Additional genotyping of genetic markers located in this specific region demonstrated significant evidence that the FE locus is situated between the chromosome 5q markers D5S642 and D5S816 (multipoint LOD score of 6.49). Notably, this region contains the cytokine gene cluster, which includes three genes-namely, those for interleukin (IL)-3, IL-5, and granulocyte/macrophage colony-stimulating factor (GM-CSF)-whose products play important roles in the development and proliferation of eosinophils. These three cytokine genes were screened for potential disease-specific mutations by resequencing of a subgroup of individuals from the present kindred. No functional sequence polymorphisms were found within the promoter, the exons, or the introns of any of these genes or within the IL-3/GM-CSF enhancer, suggesting that the primary defect in FE is not caused by a mutation in any one of these genes but, rather, is caused by another gene in the area.
Adolescent↗
Intestinal reperfusion-induced pulmonary edema is related to increased pulmonary inducible nitric oxide synthase activity.
BACKGROUND: This study examines the hypothesis that specific inhibition of the inducible isoform of nitric oxide synthase (iNOS) will attenuate intestinal reperfusion-induced pulmonary microvascular dysfunction. METHODS: Sprague-Dawley rats underwent intestinal ischemia-reperfusion (IR) or sham operation (SHAM). Before injury, the animals received a selective inhibitor of iNOS (S-methylisothiourea sulfate, SMT: L-N6-[1-iminoethyl] lysine L-NIL), a nonselective inhibitor of NOS (NG-nitro-L-arginine methylester, L-NAME) or vehicle (0.9% saline). IR-induced changes in pulmonary microvascular permeability were assessed by quantitating the extravasation of Evans blue dye (EBD)-bound protein into the lung. Pulmonary iNOS activity and content were assessed by radiochemical analysis and Western blot, respectively. RESULTS: There was 60% more EBD within the lungs of animals sustaining IR when compared with controls (P < .05). Pretreatment with SMT or L-NIL totally prevented the increase in EBD extravasation associated with IR. In contrast, pretreatment with L-NAME resulted in a 10% increase in dye extravasation in those animals sustaining IR when compared with similarly injured animals receiving saline (P > .05). There was significantly greater iNOS activity and enzyme content within the lungs of animals sustaining IR compared with controls. CONCLUSIONS: These data are consistent with the hypothesis that the release of nanomolar quantities of nitric oxide generated by iNOS contributes to IR-induced pulmonary microvascular dysfunction.
Animals↗
Inhibition of LDL oxidation by melatonin requires supraphysiologic concentrations.
Melatonin has been suggested as a potent antioxidant that may protect against development of atherosclerosis and cancer; however, these effects are unproven and controversial. The antioxidant capacity of melatonin was tested in comparison with alpha-tocopherol, ascorbic acid, and the melatonin precursors tryptophan and serotonin, by measuring inhibition of metal ion-mediated and human macrophage-mediated oxidation of LDL. Melatonin had weak antioxidant activity that was detectable only at concentrations 10000- to 100000-fold higher than physiologic concentrations. These results were comparable with published data showing that the radical scavenging activity of melatonin requires markedly supraphysiologic concentrations. In contrast, alpha-tocopherol was 50- to 100-fold more potent and was efficacious at physiologic concentrations. Ascorbic acid and tryptophan also were active at physiologic concentrations and were significantly more potent than melatonin. In summary, extremely supraphysiologic concentrations of melatonin had only weak antioxidant activity, which was surpassed by alpha-tocopherol, ascorbic acid, and tryptophan.
Antioxidants↗
A descriptive analysis of the Journal of Manipulative and Physiological Therapeutics, 1989-1996.
BACKGROUND: Two previous reports have summarized the content, institutional affiliations, academic training and funding sources for articles published in the Journal of Manipulative and Physiological Therapeutics (JMPT) from 1978-1986 and 1987-1988. OBJECTIVES: (a) to quantitatively assess the types of articles published in the JMPT from 1989-1996; (b) to identify the affiliations of contributors to the JMPT during this period; (c) to identify the academic backgrounds of contributors to the JMPT from 1989-1996; (d) to identify funding sources for scholarly works published in the JMPT during this period; (e) to identify the proportionate contributions of female authors; (f) to assess the proportion of articles contributed, i.e., foreign vs. domestic sources; and (g) to compare findings for the JMPT from 1989-1996 with similar data for 1978-1988. STUDY DESIGN: Survey of the contents of the JMPT from 1989-1996. METHODS: The contents of the 69 issues of the JMPT from 1989-1996 were reviewed by all authors. Characteristics extracted included category of the article, academic backgrounds of authors, institutional affiliations of authors, funding sources, gender of authors and nation(s) of origin of articles. RESULTS: The annual rate of published contributions to the Journal has more than doubled compared with its first 11 yr of publication, and the proportion of original data reports has grown slightly. Controlled and quasicontrolled clinical trials were 7 times more numerous (n = 28 articles) during the past 8 yr. Chiropractic colleges were the most frequently mentioned affiliation of authors, followed by private practice and nonchiropractic colleges. Collaborative articles submitted by authors at two or more chiropractic colleges grew from only 4 articles from 1978-1988 to 31 articles from 1989-1996. As in previous years, the National College of Chiropractic continued to be the most frequently mentioned academic affiliation of authors. The numbers of articles contributed by those holding scientific (e.g., PhD) and medical degrees have grown substantially. The number of articles mentioning financial support grew from 78 from 1978-1988 to 179 from 1989-1996, and 58 new funding sources were identified. The Foundation for Chiropractic Education and Research continues to be the most frequently mentioned source of funding. Of all articles published in the JMPT from 1989-1996, 21% were authored or coauthored by women. Of 1050 articles, 286 (27%) were authored or coauthored by individuals residing outside the United States of America. CONCLUSIONS: Substantial increases in scholarly activities within the chiropractic profession are suggested by the growth in scholarly products published in the discipline's most distinguished periodical. Increases in controlled outcome studies, collaboration among chiropractic institutions, contributions from nonchiropractors, contributions from nonchiropractic institutions and funding for research suggest a degree of professional maturation and growing interest in the content of the discipline.
Authorship↗
Activation of multiple growth regulatory genes following inducible expression of IRF-1 or IRF/RelA fusion proteins.
Interferon Regulatory Factor (IRF)-1 has been characterized as an important growth regulatory and immunomodulatory transcription factor. To further characterize the potential targets of IRF-1 antiproliferative activity, IRF-1 was expressed under the control of the tetracycline-inducible system in murine NIH3T3 cells. Due to their ability to mimic IRF-1 transactivator function, the regulatory potential of IRF-1/RelA and IRF-2/RelA fusion proteins consisting of the DNA binding domains of IRF-1 or IRF-2 fused to the transactivation domain of NF-kappaB RelA(p65) was also examined. Cells inducibly expressing IRF-1 or IRF/RelA in response to doxycycline treatment displayed significantly reduced growth rates compared to control cells, and inhibition of cell growth correlated directly with the level of transgene expression. Interestingly, IRF-1 and IRF/RelA expression also induced a low level of apoptosis, as detected by microscopic analyses. Furthermore, expression of the interferon inducible, double stranded RNA dependent kinase PKR was increased following IRF-1 or IRF/RelA induction. Most strikingly, induction of IRF-1 and IRF/RelA expression resulted in a significant increase in STAT1 (p91) protein and increased ISGF3 DNA binding activity, suggesting that IRF-1 tumor suppressor activity may involve a novel mechanism which activates the JAK-STAT pathway through STAT1. WAF1 levels were also constitutively elevated in IRF-1 and IRF-1/RelA cells. These studies demonstrate that inducible expression of the transactivation function of IRF-1 or IRF/RelA mediates tumor suppressor activity by inducing cell growth arrest, apoptosis and the differential activation of growth regulatory genes such as PKR, STAT1 and WAF1.
3T3 Cells↗
Tumor type and vascularity: important variables in infusional brachytherapy with colloidal 32P.
PURPOSE: This study investigated the role of histologic tumor characteristics, in comparison with a normal tissue, and of tumor vascularization on the uptake and retention of colloidal 32P used in infusional brachytherapy of solid cancers. The cytotoxicity of colloidal 32P was also evaluated for two tumors of different radiosensitivity, a melanoma, and a squamous cell carcinoma. METHODS AND MATERIALS: An in vitro analysis of colloidal 32P uptake was carried out on a human melanoma cell line, HBL, a human squamous cell carcinoma, SCC1, and normal fibroblasts, F-NBB. Tumor retention of colloidal 32P was studied in vivo for the HBL and the SCC1 tumors implanted subcutaneously in nude mice. Tumor vascular density was determined by microscopic study of Masson's trichrome slides of HBL and SCC1 tumors of about 1 cm diameter. RESULTS: In vitro studies showed that the time required for maximal cell uptake of colloidal 32P was only 10-20 min for the SCC1 and HBL tumors, while it took at least 60 min for the fibroblasts. After intratumoral injection of macroaggregated albumin (MAA), followed by 50 microCi of colloidal 32P, Bremsstrahlung imaging performed at 6 and 24 h showed that the activity remained in the HBL tumor while some of the radiocolloids leaked from the SCC1 tumor and was trapped in the reticuloendothelial system of the liver. Organ activity counting confirmed this finding: 32P activity was three to four times higher in the HBL than in the SCC1 tumor, whereas the activity in the liver, insignificant in the HBL mice (less than 0.1 microCi/g), was as high as 24 microCi/g in the SCC1 mice. This phenomenon may be explained by the difference in tumor vascular density, estimated for the HBL to be about four times less than that of the SCC1 tumor (5.7 vs. 21.4 blood vessels per mm2 for the HBL and the SCC1 tumors, respectively). CONCLUSION: Intratumoral infusion of colloidal 32P may be a useful complement of radiation therapy in the treatment of nonresectable but accessible solid tumors. Tumor vascularization must be taken into account for a successful vascular blockade by MAA prior to the infusion of colloidal 32P.
Animals↗
Enhanced ribosomal association of p27(Kip1) mRNA is a mechanism contributing to accumulation during growth arrest.
p27(Kip1) regulates the decision to enter into S-phase or withdraw from the cell cycle by establishing an inhibitory threshold above which G1 cyclin-dependent kinases accumulate before activation. We have used the HL-60 cell line to study regulation of p27 as cells withdraw from the cell cycle following treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA). We found that the amount of p27 is maximal in G0 cells, lower in G1 cells, and undetectable in S-phase cells. In contrast to the protein, the amount of p27 mRNA was the same in these populations, suggesting that accumulation of p27 during the cell cycle and as cells withdraw from the cell cycle is controlled by post-transcriptional mechanisms. In S-phase cells, the degradation of p27 appears to predominate as a regulatory mechanism. In G0 cells, there was an increase in the synthesis rate of p27. Our data demonstrate that, in G0 cells, accumulation of p27 is due to an increase in the amount of p27 mRNA in polyribosomes.
Cell Cycle↗