Synthesis of the titanium compound [(MeC(5)H(4)TiCl(mu-NSiMe(3)](2) with migration of a SiMe(3) group and preparation of CpZrCl(eta(2)-NHNCHSiMe(3)).
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Journal Article↗
Biomedical subjects
H Hao
Publications and source records attributed to H Hao.
Multiple levels of regulation of selenoprotein biosynthesis revealed from the analysis of human glioma cell lines.
To gain a better understanding of the biological consequences of the exposure of tumor cells to selenium, we evaluated the selenium-dependent responses of two selenoproteins (glutathione peroxidase and the recently characterized 15-kDa selenoprotein) in three human glioma cell lines. Protein levels, mRNA levels, and the relative distribution of the two selenocysteine tRNA isoacceptors (designated mcm(5)U and mcm(5)Um) were determined for standard as well as selenium-supplemented conditions. The human malignant glioma cell lines D54, U251, and U87 were maintained in normal or selenium-supplemented (30 nM sodium selenite) conditions. Northern blot analysis demonstrated only minor increases in steady-state GSHPx-1 mRNA in response to selenium addition. Baseline glutathione peroxidase activity was 10.7 +/- 0.7, 7.6 +/- 0.7, and 4.3 +/- 0.7 nmol NADPH oxidized/min/mg protein for D54, U251, and U87, respectively, as determined by the standard coupled spectrophotometric assay. Glutathione peroxidase activity increased in a cell line-specific manner to 19.7 +/- 1.4, 15.6 +/- 2.1, and 6. 7 +/- 0.5 nmol NADPH oxidized/min/mg protein, respectively, as did a proportional increase in cellular resistance to H(2)O(2), in response to added selenium. The 15-kDa selenoprotein mRNA levels likewise remained constant despite selenium supplementation. The selenium-dependent change in distribution between the two selenocysteine tRNA isoacceptors also occurred in a cell line-specific manner. The percentage of the methylated isoacceptor, mcm(5)Um, changed from 35.5 to 47.2 for D54, from 38.1 to 47.3 for U251, and from 49.0 to 47.6 for U87. These data represent the first time that selenium-dependent changes in selenoprotein mRNA and protein levels, as well as selenocysteine tRNA distribution, were examined in human glioma cell lines.
Gene Expression Regulation, Neoplastic↗
Study of phonon dispersion in silicon and germanium at long wavelengths using picosecond ultrasonics
We have studied the dispersion of long wavelength longitudinal phonons in silicon and germanium using ultrasonic techniques. For long wavelengths, the acoustic phonon dispersion relation is of the form omega(k) approximately ck-gammak(3), where c is the speed of sound and gamma measures the lowest-order phonon dispersion. By sending an ultrasonic pulse of length a few hundred angstroms into a crystal and measuring the change of the pulse shape with propagation distance, we are able to determine the parameter gamma. The results are compared with lattice dynamics models.
Journal Article↗
The First Structurally Characterized Metal-SeH Compounds:
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Journal Article↗
Are A3 adenosine receptors expressed in the brain?
An increasing number of reports suggest a role for A3 adenosine receptors (A3ARs) in mediating adenosine action in the central nervous system. However, studies of A3AR localization in the brain have yet to be performed. To provide insights into the central sites of A3AR action, we compared patterns of A1 and A3AR mRNA and binding site expression in the brains of rats, mice and humans. We also assessed whether A3 agonists are selective for A3ARs. Whereas it was possible to detect high-level A1AR expression in many brain regions, it was not possible to detect either A3AR gene or binding site expression in the central nervous system. When we examined the affinities of the A3AR agonists CI-IAB-MECA and IAB-MECA for A1ARs, we found that these compounds bound to A1ARs with high affinity. These observations suggest that studies using A3-agonists need to consider potential effects of A1ARs activation, as A1ARs are abundantly expressed in the nervous system whereas A3AR expression in the brain cannot be directly demonstrated.
Aged↗
Knowledge-based grouping of modeled HLA peptide complexes.
Human leukocyte antigens are the most polymorphic of human genes and multiple sequence alignment shows that such polymorphisms are clustered in the functional peptide binding domains. Because of such polymorphism among the peptide binding residues, the prediction of peptides that bind to specific HLA molecules is very difficult. In recent years two different types of computer based prediction methods have been developed and both the methods have their own advantages and disadvantages. The nonavailability of allele specific binding data restricts the use of knowledge-based prediction methods for a wide range of HLA alleles. Alternatively, the modeling scheme appears to be a promising predictive tool for the selection of peptides that bind to specific HLA molecules. The scoring of the modeled HLA-peptide complexes is a major concern. The use of knowledge based rules (van der Waals clashes and solvent exposed hydrophobic residues) to distinguish binders from nonbinders is applied in the present study. The rules based on (1) number of observed atomic clashes between the modeled peptide and the HLA structure, and (2) number of solvent exposed hydrophobic residues on the modeled peptide effectively discriminate experimentally known binders from poor/nonbinders. Solved crystal complexes show no vdW Clash (vdWC) in 95% cases and no solvent exposed hydrophobic peptide residues (SEHPR) were seen in 86% cases. In our attempt to compare experimental binding data with the predicted scores by this scoring scheme, 77% of the peptides are correctly grouped as good binders with a sensitivity of 71%.
Alleles↗
Developing circadian rhythmicity.
Circadian rhythms are endogenously generated rhythms with a period length of about 24-hours. Evidence gathered over the past decade indicates that the circadian timing system develops prenatally, and the suprachiasmatic nuclei, the site of a circadian clock, is present by midgestation in human and nonhuman primates. Recent evidence also shows that the circadian system of primate infants is responsive to light at very premature stages and that low intensity lighting can regulate the developing clock. After birth, there is progressive maturation of the circadian system outputs, with pronounced rhythms in sleep-wake and hormone secretion generally developing after 2 months of age. With the continued elucidation of circadian system development and influences on human physiology and illness, it is anticipated that consideration of circadian biology will become an increasingly important component of neonatal care.
Animals↗
Specific sequences outside the E-box are required for proper per expression and behavioral rescue.
A 69 bp circadian regulatory sequence (CRS) upstream of the per gene is sufficient to drive circadian transcription, mediate proper spatial expression, and rescue behavioral rhythmicity in per01 flies. Within the CRS, an E-box is required for transcriptional activation by two basic-helix-loop-helix (bHLH) PERARNT-SIM (PAS) transcription factors, dCLOCK (dCLK) and CYCLE (CYC). To define sequences within the CRS that are required for spatial expression, circadian expression, and behavioral rhythmicity, a series of mutants that alter blocks of 3 to 12 nucleotides across the entire CRS were used to drive lacZ or per expression in vivo. As expected, the E-box within the CRS is necessary for high-level expression and behavioral rhythmicity, but sequences outside the E-box are also required for transcriptional activation, proper spatial expression, and behavioral rhythmicity. These results indicate that the dCLK-CYC target site extends beyond the E-box and that factors other than dCLK and CYC modulate per transcription.
ARNTL Transcription Factors↗
Melatonin does not shift circadian phase in baboons.
It has been suggested that the pineal hormone melatonin can modulate circadian rhythmicity and may have clinical utility in treating biological clock disorders. Thus, there is considerable clinical interest in using melatonin to treat disorders, such as jet-lag. Yet, despite growing enthusiasm for the use of melatonin, it is not clear whether melatonin indeed shifts the circadian phase in humans and other primate species. Thus, to assess whether melatonin can influence circadian phase, we studied the phase-shifting effects of melatonin on baboons to provide insights into the role of melatonin. Mel- atonin was administered orally to baboons (0.5, 3, 5, or 10 mg) either in the early morning hours from circadian time (CT) 0 to CT3 or late in the afternoon from CT9 to CT12, and changes in circadian phase were assessed. Surprisingly, at all doses and times tested, melatonin did not shift circadian phase. Physical activity was reduced after 5- and 10-mg doses given late in the afternoon, but not after doses given early in the morning. These observations suggest that melatonin does not shift circadian phase in baboons using doses similar to those prescribed for treating human circadian system disorders.
Animals↗
Bleomycin-induced pulmonary fibrosis is independent of eosinophils.
Eosinophils have been shown to increase in tissues during many fibrotic conditions and consequently have been suggested to contribute to the development of fibrosis. This study tested the hypothesis that eosinophils are essential in the development of lung fibrosis in mice in response to bleomycin (BLM). Anti-IL-5 antibody was administered intraperitoneally into mice 2 h prior to endotracheal BLM inoculation and thereafter, every other day. Lung eosinophilia was evaluated by measurement of eosinophil peroxidase activity and confirmed by eosinophil counts in histologic sections. Lung fibrosis was evaluated by hydroxyproline content and confirmed by collagen staining in histological sections. Results demonstrated that BLM induced pronounced lung eosinophilia, which was maximal 7 days after BLM treatment and remained elevated through day 14, in C57B1/6 SCID mice and CBA/J mice. In contrast, eosinophilia was a minor component in the lungs of wildtype C57B1/6 mice after BLM treatment, although lung fibrosis developed similarly in all three strains of mice. Treatment with anti-IL-5 completely abrogated eosinophilia but failed to block pulmonary fibrosis induced by BLM in all mouse strains, including C57B1/6 SCID, wildtype C57B1/6 mice, and CBA/J mice. Analysis of cytokine mRNA by RNase-protection assay in C57B1/6 SCID mice indicated that BLM treatment caused enhanced expression of the cytokines, TNF-alpha, and IL-6 at days 3, 7, and 14 post-BLM inoculation, regardless of whether eosinophils were depleted by anti-IL-5. Finally, the importance of eosinophils in lung fibrosis was examined in IL-5 gene knockout mice (IL-5tm1Kopf). BLM treatment induced significant lung fibrosis in IL-5 knockout mice in the absence of eosinophilia. These findings indicate that eosinophils are not an absolute requirement for BLM-induced pulmonary fibrosis in the mouse.
Animals↗
The biological clock of very premature primate infants is responsive to light.
Each year more than 250,000 infants in the United States are exposed to artificial lighting in hospital nurseries with little consideration given to environmental lighting cycles. Essential in determining whether environmental lighting cycles need to be considered in hospital nurseries is identifying when the infant's endogenous circadian clock becomes responsive to light. Using a non-human primate model of the developing human, we examined when the circadian clock, located in the hypothalamic suprachiasmatic nuclei (SCN), becomes responsive to light. Preterm infant baboons of different ages were exposed to light (5,000 lux) at night, and then changes in SCN metabolic activity and gene expression were assessed. After exposure to bright light at night, robust increases in SCN metabolic activity and gene expression were seen at ages that were equivalent to human infants at 24 weeks after conception. These data provide direct evidence that the biological clock of very premature primate infants is responsive to light.
Animals↗
The 69 bp circadian regulatory sequence (CRS) mediates per-like developmental, spatial, and circadian expression and behavioral rescue in Drosophila.
The period (per) gene is an essential component of the circadian timekeeping mechanism in Drosophila. This gene is expressed in a circadian manner, giving rise to a protein that feeds-back to regulate its own transcription. A 69 bp clock regulatory sequence (CRS) has been identified previously upstream of the period gene. The CRS confers wild-type mRNA cycling when used to drive a lacZ reporter gene in transgenic flies. To determine whether the CRS also mediates proper developmental and spatial expression and behavioral rescue, we used the CRS to drive either lacZ or per in transgenic flies. The results show that the CRS is able to activate expression in pacemaker neuron precursors in larvae and essentially all tissues that normally express per in pupae and adults. The CRS is sufficient to rescue circadian feedback loop function and behavioral rhythms in per01 flies. However, the period of locomotor activity rhythms shortens if a stronger basal promoter is used. This study shows that regulatory elements sufficient for clock-dependent and tissue-specific per expression in larvae, pupae, and adults are present in the CRS and that the period of adult locomotor activity rhythms is dependent, in part, on the overall level of per transcripts.
Animals↗
T cell independence of bleomycin-induced pulmonary fibrosis.
The role of T cells and cytokines in bleomycin (BLM)-induced fibrosis was evaluated in susceptible and resistant strains of normal and SCID mice. Histology and hydroxyproline analysis showed that BLM induced pulmonary fibrosis in C57BL/6 and (C57BL/6 x BALB/c)F1 mice, whereas BALB/c mice were resistant to the disease. To test whether lymphocytes were required for the induction of BLM-induced pulmonary fibrosis, SCID mice were injected intratracheally with BLM and evaluated for the development of pulmonary inflammation and fibrosis. Similar morphological changes and increases in hydroxyproline were observed in both C57BL/6 SCID and (C57BL/6 x CB.17)F1 SCID animals compared to those seen in wild-type C57BL/6 and (C57BL/6 x BALB/c)F1 mice. In contrast, CB.17 SCID mice, which are genetically similar to BALB/c mice, were resistant to disease induction. Analysis of the cellular infiltrate in BLM-treated C57Bl/6 SCID mice confirmed a lack of T cells in the lungs of SCID mice and demonstrated a pronounced accumulation of eosinophils in areas of developing pulmonary fibrosis. NK cells were significantly elevated in untreated SCID mice and did not increase further after BLM treatment. Analysis of selected cytokines 1 day after initiation of BLM-induced pulmonary fibrosis indicated that the levels of TNF-alpha and IFN-gamma appeared to segregate with fibrosis in both the SCID and wild-type mice. The data demonstrate that T cells are not required for the induction of fibrosis by BLM and suggest that responses by non-lymphoid cells may be sufficient for the induction of fibrosis.
Animals↗
Mechanisms of human mitochondrial DNA maintenance: the determining role of primary sequence and length over function.
Although the regulation of mitochondrial DNA (mtDNA) copy number is performed by nuclear-coded factors, very little is known about the mechanisms controlling this process. We attempted to introduce nonhuman ape mtDNA into human cells harboring either no mtDNA or mutated mtDNAs (partial deletion and tRNA gene point mutation). Unexpectedly, only cells containing no mtDNA could be repopulated with nonhuman ape mtDNA. Cells containing a defective human mtDNA did not incorporate or maintain ape mtDNA and therefore died under selection for oxidative phosphorylation function. On the other hand, foreign human mtDNA was readily incorporated and maintained in these cells. The suicidal preference for self-mtDNA showed that functional parameters associated with oxidative phosphorylation are less relevant to mtDNA maintenance and copy number control than recognition of mtDNA self-determinants. Non-self-mtDNA could not be maintained into cells with mtDNA even if no selection for oxidative phosphorylation was applied. The repopulation kinetics of several mtDNA forms after severe depletion by ethidium bromide treatment showed that replication and maintenance of mtDNA in human cells are highly dependent on molecular features, because partially deleted mtDNA molecules repopulated cells significantly faster than full-length mtDNA. Taken together, our results suggest that mtDNA copy number may be controlled by competition for limiting levels of trans-acting factors that recognize primarily mtDNA molecular features. In agreement with this hypothesis, marked variations in mtDNA levels did not affect the transcription of nuclear-coded factors involved in mtDNA replication.
Animals↗
Suppression of a mitochondrial tRNA gene mutation phenotype associated with changes in the nuclear background.
We previously have characterized a pathogenic mtDNA mutation in the tRNAAsn gene. This mutation (G5703A) was associated with a severe mitochondrial protein synthesis defect and a reduction in steady-state levels of tRNAAsn. We now show that, although transmitochondrial cybrids harboring homoplasmic levels of the mutation do not survive in galactose medium, several galactose-resistant clones could be obtained. These cell lines had restored oxidative phosphorylation function and 2-fold higher steady-state levels of tRNAAsn when compared with the parental mutant cell line. The revertant lines contained apparently homoplasmic levels of the mutation and no other detectable alteration in the tRNAAsn gene. To investigate the origin of the suppression, we transferred mtDNA from the revertants (143B/206 TK-) to a different nuclear background (143B/207 TK-, 8AGr). These new transmitochondrial cybrids became defective once again in oxidative phosphorylation and regained galactose sensitivity. However, galactose-resistant clones could also be obtained by growing the 8AGr transmitochondrial cybrids under selection. Because the original rate of reversion was higher than that expected by a classic second site nuclear mutation, and because of the aneuploid features of these cell lines, we searched for the presence of chromosomal alterations that could be associated with the revertant phenotype. These studies, however, did not reveal any gross changes. Our results suggest that modulation of the dosage or expression of unknown nuclear-coded factor(s) can compensate for a pathogenic mitochondrial tRNA gene mutation, suggesting new strategies for therapeutic intervention.
Animals↗
Brain natriuretic peptide enhances the endothelium-independent cAMP and vasorelaxant responses of calcitonin gene-related peptide in rat aorta.
Calcitonin gene-related peptide (CGRP) causes vasorelaxation in rat aorta involving endothelium/nitric oxide (NO)-dependent elevations of both cAMP and cGMP levels. When endothelium is removed, preincubation with exogenous NO uncovers and potentiates direct (endothelium-independent) cAMP elevations and vasorelaxations caused by CGRP. This enhancing effect of NO potentially involves elevation of cGMP and inhibition of Type III (cGMP-inhibitable) phosphodiesterase, causing accumulation of cAMP. However, NO may have other actions. The aim of the present study was to determine if brain natriuretic peptide (BNP), which elevates cGMP levels independent of NO, could enhance cAMP accumulations and vasorelaxations induced by CGRP in rat aortic rings denuded of endothelium. When added separately, neither CGRP (100 nM) nor BNP (10 nM) altered cAMP levels. When added in combination, CGRP (100 nM) and BNP (10 nM) significantly elevated cAMP levels (from control of 0.95+/-0.08 to 1.53+/-0.09 pmol/mg protein) at 2 min. BNP (10 nM) elevated cGMP levels 10-fold at 2 min and this response was not altered by co-administration of CGRP (100 nM). Pretreatment with BNP at concentrations as low as 1 nM in endothelium-denuded aortic rings greatly enhanced the direct vasorelaxant effects of CGRP (100 nM) (from control of 0% to 57.6+/-6.8% relaxation of phenylephrine-precontractions). Our findings indicate that BNP enhances direct (endothelium-independent) cAMP elevations and vasorelaxations caused by CGRP in rat aorta, thus supporting the concept that cGMP inhibits cAMP metabolism and enhances CGRP-induced responses in aortic smooth muscle cells.
Animals↗
[Pathogenesis of acute coronary syndromes].
Coronary artery diseases may categorized into asymptomatic disease, angina pectoris, myocardial infarction, chronic heart failure, and sudden coronary death. Unstable angina, acute myocardial infarction, and sudden cardiac death are known as the acute coronary syndromes. Coronary atheroma is unstable in the patients with acute coronary syndromes. Stable plaques will be unstable when dynamic alterations occur. The alterations are plaque rupture, plaque hemorrhage, coronary thrombosis and vasospasm. They act each other. We analysed the histopathology of coronary arteries who died of acute myocardial infarction in 85 cases. It showed that the risk factors of plaque rupture are clusters of form cells, eccentric plaque with soft lipid rich core, and thinning of fibrous cap in atheroma. Most of these cases ruptured at edge of the atheroma.
Acute Disease↗
Functional and structural features of a tandem duplication of the human mtDNA promoter region.
An approximately 260-bp tandem duplication of the human mtDNA regulatory region has been identified in patients with mitochondrial disorders and in a specific Caucasian haplogroup. The functional significance of this mtDNA duplication was difficult to assess, because it was present at very low levels in human tissues. We have isolated several transmitochondrial cybrid lines harboring this mutation, one of which (clone CA17.1) was essentially homoplasmic for the duplication. Oxidative-phosphorylation function was not impaired in clone CA17.1, suggesting that this mtDNA alteration is not pathogenic. mtDNA copy number and steady-state levels of heavy- and light-strand transcripts were unaltered in clone CA 17.1. The steady-state levels of RNAs made from the two promoters (either from the heavy-strand or from the light-strand) were also similar, indicating that oppositely oriented promoters did not interfere with each other.
Base Sequence↗