Structural and solution calorimetric studies of sulfur binding to nucleophilic carbenes.
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Journal Article↗
Biomedical subjects
J Huang
Publications and source records attributed to J Huang.
Bovine NAD+-dependent isocitrate dehydrogenase: alternative splicing and tissue-dependent expression of subunit 1.
NAD+-dependent isocitrate dehydrogenase (IDH), a key regulatory enzyme in the Krebs cycle, is a multi-tetrameric enzyme. At least three of the subunits in the core tetramer of mammals are unique gene products. Subunits 1/beta and 2/gamma are considered to be regulatory, while subunits 3,4/alpha, comprising half the tetramer, are catalytic. The full sequence was obtained for the major subunit 1 cDNA in bovine heart, IDH 1-A. A second cDNA, rare in heart, was also identified (IDH 1-B). Differences in the two were confined to the 3'-region, suggesting alternative splicing. Screening of brain, kidney, and liver RNA showed the presence of IDH 1-A and 1-B and a third major species, IDH 1-C. Amplification of bovine genomic DNA by PCR across the regions of difference produced a single product. Comparison of the genomic and mRNA sequences showed that IDH 1-A resulted from splicing of exon W to exon Y, eliminating intron w, exon X, and intron x. IDH 1-B was formed by splice junctions between exon W, exon X, and exon Y. IDH 1-C resulted from splicing of exon W to exon X and subsequent retention of intron x. The 2 proteins predicted from these 3 mRNAs are identical over their first 357 residues. Protein IDH 1-A, resulting from a termination codon within exon Y, contains an additional 26 residues. Proteins IDH 1-B and 1-C derive from a common termination codon within exon X and contain an additional 28 residues. The two C-terminal regions differ notably in the number and nature of charged residues, resulting in proteins with a charge difference of 3.2 at pH 7.0. Subunit 1 sequences previously reported from other species grouped with one or the other of the bovine proteins. No evidence was found for alternative splicing in subunit 3,4/alpha. The results of the present study, together with recent work on the 2/gamma subunit [Brenner,V., Nyakatura, G., Rosenthal, A., and Platzer, M. (1998) Genomics 44, 8], indicate that the regulatory subunits of the enzyme, but not the catalytic, possess alternatively spliced forms varying in C-terminal properties with tissue-specific expression. The finding is suggestive of a mechanism for modulation of allosteric regulation tailored to the needs of different tissues.
Alternative Splicing↗
The production of recombinant proteins in transgenic barley grains.
The grain of the self-pollinating diploid barley species offers two modes of producing recombinant enzymes or other proteins. One uses the promoters of genes with aleurone-specific expression during germination and the signal peptide code for export of the protein into the endosperm. The other uses promoters of the structural genes for storage proteins deposited in the developing endosperm. Production of a protein-engineered thermotolerant (1, 3-1, 4)-beta-glucanase with the D hordein gene (Hor3-1) promoter during endosperm development was analyzed in transgenic plants with four different constructs. High expression of the enzyme and its activity in the endosperm of the mature grain required codon optimization to a C+G content of 63% and synthesis as a precursor with a signal peptide for transport through the endoplasmic reticulum and targeting into the storage vacuoles. Synthesis of the recombinant enzyme in the aleurone of germinating transgenic grain with an alpha-amylase promoter and the code for the export signal peptide yielded approximately 1 microgram small middle dotmg(-1) soluble protein, whereas 54 microgram small middle dotmg(-1) soluble protein was produced on average in the maturing grain of 10 transgenic lines with the vector containing the gene for the (1, 3-1, 4)-beta-glucanase under the control of the Hor3-1 promoter.
Blotting, Southern↗
Chromatographic behavior of mouse serum immunoglobulin G in protein G perfusion affinity chromatography.
In this study chromatographic behavior of mouse serum immunoglobulin G (IgG) on a protein G perfusion affinity chromatographic column was investigated experimentally. The results indicate that the protein G column has no non-specific binding to the other proteins in mouse serum but an irreversible adsorption to IgG under the conditions investigated. It was found that variations of the elution solution composition, ionic strength and pH played to some extent an essential effect on the chromatographic behavior of IgG. The influence of the mobile phase flow-rate on the chromatographic behavior of IgG was also researched. These results show that the dissociation of IgG from protein G affinity packings becomes the rate-limiting step in the perfusion affinity chromatographic separation process.
Adsorption↗
Asbestos content in the lymph nodes of nonoccupationally exposed individuals.
BACKGROUND: The thoracic lymph nodes are a part of the clearance system from lung tissue. Accumulation of dust in these nodes are known to occur following some types of exposure. However, no information exists as to asbestos content in lymph nodes from the general population. METHODS: The study cohort consisted of 21 individuals previously defined as nonoccupationally exposed to asbestos. Tissue burden of asbestos obtained from lung analysis by analytical electron microscopy was compared with burden in the lymph nodes. RESULTS: No asbestos fibers were detected in nodes from 8 cases. The majority of the fibers found in lymph nodes were short (<5 microm) and most often noncommercial amphiboles. Ferruginous bodies (FBs) were detected in lymph node from only two samples. CONCLUSIONS: The total asbestos burden in the lung tissue from these individuals was quite low. However, in 12 of the 13 cases that had positive nodes, the tissue burden in the node was appreciably heavier per gram than in the lung. This raises the question as to whether the lymph nodes, though less efficient clearance, may be better indicators of lifetime exposure to dust than lung tissue.
Adolescent↗
Effect of allelic heterogeneity on the power of the transmission disequilibrium test.
Due in part to an influential paper by Risch and Merikangas [(1996) Science 273:1516-1517], which suggested that disequilibrium tests would have greater power to detect genes of small effect than would linkage tests, interest in the use of the Transmission Disequilibrium Test (TDT) as an analysis tool for genomewide studies is steadily growing. However, the paper by Risch and Merikangas made several simplifying assumptions. One such assumption was that the underlying gene showed allelic homogeneity, and another was that the allele being measured was the actual susceptibility allele. Here we investigate the effect of allelic heterogeneity on the power of the TDT using multiplicative, additive, dominant, and recessive modes of inheritances in the context of a genomewide study. We further distinguish two cases: first, that the marker alleles are the actual susceptibility alleles, and second, that alleles are measured at a marker linked to the disease gene with zero recombination. We consider two family structures, either a single affected offspring (SAO) and two parents, or an affected sib-pair (ASP) and two parents. We find that, as expected, the power of the TDT declines as the number of susceptibility alleles at the locus being tested increases and the effect on power can be substantial. When a linked marker is measured rather than a susceptibility allele itself, sample sizes reach unattainable levels when as few as two susceptibility alleles are present. Across all the models we consider, the required number of families for a TDT with ASP sampling varies from 19 to over a million families. Thus, the TDT may not be an optimal test in the context of genomic screens under more biologically realistic assumptions.
Alleles↗
Murine Myak, a member of a family of yeast YAK1-related genes, is highly expressed in hormonally modulated epithelia in the reproductive system and in the embryonic central nervous system.
We have cloned a mouse homologue (designated Myak) of the yeast protein kinase YAK1. The 1210 aa open reading frame contains a putative protein kinase domain, nuclear localization sequences and PEST sequences. Myak appears to be a member of a growing family of YAK1-related genes that include Drosophila and human Minibrain as well as a recently identified rat gene ANPK that encode a steroid hormone receptor interacting protein. RNA blot analysis revealed that Myak is expressed at low levels ubiquitously but at high levels in reproductive tissues, including testis, epididymis, ovary, uterus, and mammary gland, as well as in brain and kidney. In situ hybridization analysis on selected tissues revealed that Myak is particularly abundant in the hormonally modulated epithelia of the epididymis, mammary gland, and uterus, in round spermatids in the testis, and in the corpora lutea in the ovary. Myak is also highly expressed in the aqueduct of the adult brain and in the brain and spinal cord of day 12.5 embryos.
Animals↗
Regional hyperthermia in the treatment of primary hepatic carcinoma.
BACKGROUND AND OBJECTIVES: The purpose of this research was to investigate the clinical significance of regional hyperthermia in the treatment of primary hepatic carcinoma (PHC). METHODS: The regional hyperthermia (60 degrees C) was used whenever there was suspicion of residual cancer tissues on the edge of the hepatic resection. The hyperthermia was maintained for about 5-20 min depending on the size and the amounts of residual nodulus. If there was obvious necrosis on the hyperthermia-treated site, the solidified tissues were removed. Otherwise, the solidified tissues were kept in situ. RESULTS: There were 68 cases of PHC patients in this study. The patients were divided into 4 groups: A, lobectomy, 14 cases; B, lobectomy plus regional hyperthermia, 12 cases; C, regional hepatectomy, 16 cases; D, regional hepatectomy plus regional hyperthermia, 26 cases. All patients were followed after their operations. The mean survival time of the 4 groups was as follows: Group A, 346.5 days (186-921 days); Group B, 432.6 days (254-1189 days); Group C, 525.4 days (192-1016 days); and Group D, 1142 (from 318 days to seven years and two months) days. There were significant differences between Groups A and B (P < 0.01) and between Groups C and D (P < 0.01). It seems that regional hyperthermia on the hepatic resection edge can prolong patients' survival time. CONCLUSIONS: Regional hyperthermia on the hepatic resection edge is helpful for prolonging patients' survival time in the treatment of primary hepatic carcinoma.
Adult↗
TCR antagonist peptides induce formation of APC-T cell conjugates and activate a Rac signaling pathway.
T cell receptor antagonists inhibit T cell activation by antigen, and by themselves fail to induce phenotypic changes associated with T cell activation. However, they can induce limited tyrosine phosphorylation of TCRzeta chain. Here we show that TCR antagonists are potent inducers of APC-T cell conjugates, cytoskeletal reorganization, and capping of certain T cell proteins. These events are associated with a signaling pathway involving tyrosine phosphorylation of Vav and SLP-76, activation and capping of Rac-1, a protein previously linked with cytoskeletal reorganization, and activation of JNK. The finding that antagonist peptides stimulate this pathway, while failing to stimulate other TCR-mediated signaling pathways, indicates the presence in T cells of a hierarchy of signaling that is sensitive to the avidity of Ag / MHC-TCR interaction.
Adaptor Proteins, Signal Transducing↗
Correlation of coronary angiography with "tombstoning" electrocardiographic pattern in patients after acute myocardial infarction.
BACKGROUND: It has been suggested that a specific pattern of electrocardiographic (ECG) changes following acute myocardial infarction (AMI), the so-called "tombstoning," predicts a poorer outcome, although the cause and associations of such changes are not known. To address the possible cause and implications, we correlated angiographic findings with tombstoning in patients following AMI. METHODS: The study investigated 124 patients with AMI, whose ECGs were taken within 24 h of onset of symptoms and who subsequently underwent angiography. In this population, 24 (19%) patients had a definite tombstoning pattern on their admission ECG. RESULTS: Compared with non-tombstoning ECGs, the significant differences in the tombstoning group are as follows: (1) All patients, including those with inferior infarction, had either total or partial occlusion of the left anterior descending (LAD) artery (100 vs. 44%, p < 0.0001); (2) LAD occlusions were significantly more severe and mostly proximal (100% occlusion: 50 vs. 20.5%, p = 0.02; <50% occlusion: 0 vs. 15.9% p = 0.039; proximal occlusion: 92 vs. 65%, p = 0.017); (3) patients with tombstoning ECGs had a significantly greater incidence of occlusion of all three coronary arteries (54.1 vs. 22%, p = 0.001); (4) tombstoning ECGs were more strongly associated with anterior than with inferior infarction (83.3 vs. 33%, p < 0.0001). CONCLUSIONS: The patients with a tombstoning pattern on the admission ECG, who underwent angiography, were associated with occlusion of a high-grade stenosis of the proximal LAD artery (usually with involvement of more than one artery) and were predominantly seen in association with anterior infarction.
Adult↗
Optical coherence tomography (OCT) abnormalities in rhodopsin mutant transgenic swine with retinal degeneration.
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Animals↗
Expression of alpha-expansin genes in young seedlings of rice (Oryza sativa L.).
Rice is the only cereal in which germination and coleoptile elongation occur in hypoxia or anoxia. Little is known of the molecular basis directly underlying coleoptile cell extension. In this paper, we describe the expression of alpha-expansin genes in embryos during seed development and young seedlings grown under various oxygen concentrations. The genes Os-EXP2 and Os-EXP1 were predominantly expressed in the developing seeds, mainly in newly developed leaves, coleoptiles, and seminal roots. These expansins expressed in the developing seeds may give cells the potential to expand after seed imbibition begins. In coleoptiles, Os-EXP4 and Os-EXP2 mRNAs were greatly induced by submergence, while they were weakly detected in aerobic or anoxic conditions. Under submerged soil conditions, the signals hybridized with probes Os-EXP4 and Os-EXP2 in coleoptiles were strongest when coleoptiles elongated in the water layer. These data show that expansin gene expression is highly correlated with coleoptile elongation in response to oxygen concentrations. The Os-EXP4 gene was also expressed in leaves, mesocotyls, and coleorhizas of young seedlings. The growth of these tissues was also correlated with the presence of expansins. Therefore, the evidence derived from this study clearly demonstrates that expansins are indispensable for the growing tissues of rice seedlings.
Gene Expression Regulation, Developmental↗
A novel association between the human heat shock transcription factor 1 (HSF1) and prostate adenocarcinoma.
A search for differentially expressed genes in a pair of nonmetastatic (PC-3) versus metastatic variant (PC-3M) human prostate carcinoma cell lines led to identification of the human heat shock factor (HSF1) as an overexpressed gene product in PC-3M cells. Analysis of primary prostate cancer specimens indicated that HSF1 is generally up-regulated in most of the malignant prostate epithelial cells relative to the normal prostate cells. Among the known effectors of HSF1 action, constitutive levels of HSP70 and HSP90 are not significantly altered by the naturally elevated expression of HSF1 as in PC-3M cells or by transduced overexpression of HSF1 in PC-3 cells. The basal levels of HSP27 in both cases are, however, consistently increased by two- to threefold. With respect to response to heat shock, high basal concentration of HSP90 is not further enhanced in these cells, and HSP70 is up-regulated irrespective of HSF1 level. Heat shock, however, causes an increase in HSP27 when HSF1 is up-regulated, except when the expression of HSF1 is already too high. These results document for the first time that HSF1 is overexpressed in human prostate cancer cells, at least one consequence of which in the prostate cancer cell lines tested is stimulation of both basal and stress-induced expression of HSP27, an important factor in cell growth, differentiation, or apoptosis.
Adenocarcinoma↗
Inhibitors of HMG-CoA reductase sensitize human smooth muscle cells to Fas-ligand and cytokine-induced cell death.
Hydroxymethylglutaryl CoA (HMG CoA) reductase inhibitors, or statins, have been shown to reduce atherosclerotic cardiovascular morbidity and mortality. Atherosclerotic plaque lesions can be chronically inflamed and vulnerable to rupture or stable and less rupture-prone. Human smooth muscle cells (SMC) are critically important in maintaining the stability of atherosclerotic plaques. This stability may be greatly influenced by pro-inflammatory mediators such as IFN-gamma, TNF-alpha, and Il-1beta and Fas ligand (FasL) that are present in human atheroma. The purpose of the present study was to examine the effect of the statins on apoptosis of SMC. We have found that SMC are normally resistant to Fas or cytokine-induced apoptosis, but can be sensitized to these agents with pharmacological concentrations of some statins. Simvastatin and lovastatin strongly sensitized the cells to apoptotic agents while atorvastatin was less effective. In contrast to the lipophilic statins, the hydrophilic statin pravastatin did not induce this sensitization of SMC to apoptosis. Treatment of SMC with either mevalonate, the product of the HMG-CoA reductase, or geranylgeranylpyrophosphate, a down stream intermediate, prevented lipophilic statin-induced sensitization to apoptosis. These results suggest that prenylation of one or more proteins is critically involved in regulating the sensitivity of SMC to apoptotic stimuli. Our data support the emerging evidence that through this pathway the various statins may have effects which are beyond a simple lowering of the levels of circulating cholesterol.
Apoptosis↗
Anti-VEGF antibody suppresses primary tumor growth and metastasis in an experimental model of Wilms' tumor.
BACKGROUND/PURPOSE: Vascular endothelial growth factor (VEGF) has been shown previously to correlate with tumor growth and metastasis in an experimental model of anaplastic Wilms' tumor. The authors hypothesized that treatment with anti-VEGF antibodies would suppress both primary tumor growth and metastasis in this model. METHODS: Tumors were induced in the right kidneys of nude mice by the injection of cultured Wilms' tumor cells. After 1 week, anti-VEGF treatment was begun with injection of either vehicle or an anti-VEGF antibody intraperitoneally. Mice were killed after 4.5 weeks of treatment and tumor weights and the incidence of metastases evaluated. RESULTS: Anti-VEGF treatment resulted in a greater than 95% reduction in tumor weight (P < .0001). Anti-VEGF treatment also abolished the establishment of lung metastases (40% in control animals, P < .003). Cessation of treatment resulted in rebound tumor growth. CONCLUSION: Anti-VEGF therapy can suppress both primary tumor growth and the establishment of metastases in experimental anaplastic Wilms' tumor.
Animals↗
Triterpenoid saponins from Ardisia mamillata.
Two saponins were isolated from the roots of Ardisia mamillata HANCE. Their structures were established on the basis of MALDI-TOFMS, 1H, 13C NMR and 2D NMR (COSY, HOHAHA, HETCOR, HMBC and ROESY) spectra, and on chemical evidence, to be ardisimamilloside A, 3-O-[alpha-L-rhamnopyranosyl-(1 --> 2)-beta-D-glucopyranosyl-(1 --> 4)-[beta-D-glucopyranosyl-(1 --> 2)]-alpha-L-arabinopyranosyl]-3beta, 16alpha,28alpha-trihydroxy-13beta,28-epoxy-oleanan+ ++-30-al; and ardisimamilloside B, 3-O-[alpha-L-rhamnopyranosyl-(1 --> 2)-beta-D-glucopyranosyl-(1 --> 4)-[beta-D-glucopyranosyl-( 1 --> 2)]-alpha-L-arabinopyranosyl]-beta3-hydroxy-13beta,28- epoxy-oleanan-16-oxo-30-al.
Carbohydrate Conformation↗
Synergistic effect of Tripterygium wilfordii Hook.F (TWHF) and cyclosporin A in rat liver transplantation.
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Animals↗
The role of carbonate radical in limiting the persistence of sulfur-containing chemicals in sunlit natural waters.
Carbonate radical (*CO3-) is a selective oxidant that may be important in limiting the persistence of a number of sulfur-containing compounds in sunlit natural waters. Thioanisole, dibenzothiophene (DBT), and fenthion were selected to investigate the degradation pathway initiated by *CO3-; electron-rich sulfur compounds are particularly reactive towards the *CO3-. Using HPLC, GC, GC-MS and LC-MS for structural confirmation, the major photodegradation products of thioanisole and DBT were the corresponding sulfoxides. The sulfoxide products were further oxidized through reaction with *CO3- to the corresponding sulfone derivatives. Fenthion showed a similar pathway with appearance of fenthion sulfoxide as the major product. The proposed mechanism involves abstraction of an electron on sulfur to form a radical cation, which is then oxidized by dissolved oxygen. Each of the sulfur probes were further investigated in a sunlight simulator under varying matrix conditions. The highest rate constants occurred in the *CO3- matrix, and the lowest occurred in a matrix of dissolved organic carbon (DOC) and bicarbonate. In synthetic and natural field water, thioanisole photodegraded faster than under direct photolysis, with half-lives of 75.1 and 85.8 min, respectively. Fenthion photodegraded more rapidly than thioanisole. DBT photodegraded rapidly in a *CO3- matrix with a half-life of 24.8 min, while the half-life of direct photolysis was 350 min. Photodegradation products of each compound were also investigated. Ultimately, *CO3- was found to contribute toward the photodegradation of sulfur-containing compounds in natural waters.
Carbonates↗