Dihydropyrimidine dehydrogenase activity and fluorouracil chemotherapy.
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Dihydrouracil Dehydrogenase (NADP)↗
Biomedical subjects
Z Lu
Publications and source records attributed to Z Lu.
Characterization of the translocation process of vitamin D3 from the skin into the circulation.
The cutaneous synthesis of vitamin D3 and the subsequent translocation of vitamin D3 into the circulation are two key steps in the vitamin D endocrine system. To study the kinetic aspects of cutaneous synthesis and translocation of vitamin D3, both in vitro and in vivo chicken models have been developed. To assess the capacity of chicken skin to generate vitamin D3, the concentrations of 7-dehydrocholesterol (7-DHC) in different skin areas were determined. It was found that the highest concentration of 7-DHC was in the leg skin (3524 +/- 937 ng cm-2), which was about 30 times greater than that in the back (120 +/- 62 ng cm-2). Whole body exposure of chickens to UV-B radiation (0.5 J cm-2) resulted in the production of previtamin D3 (preD3) in the skin of the legs and feet (43 +/- 7 and 54 +/- 17 ng cm-2, respectively), whereas no preD3 was detected in the back skin. In vitro, at 40 C, the forward (k1) and reverse (k2) rate constants of the preD3<-->vitamin D3 reaction in the leg skin were greatly increased compared to those in n-hexane (k1, 0.367 vs. 0.0369 h-1; k2, 0.042 vs. 0.0059 h-1). In vivo, the determined rate constants k1, k2, and k3 for the consecutive reactions preD3<-->vitamin D3-->vitamin D3 were 0.257, 0.034, and 0.114 h-1, respectively. To evaluate the circulating concentration of vitamin D3 in response to UV-B radiation, chicken legs were irradiated. The time course revealed a 4-fold increase in the circulating concentration of vitamin D3, with a peak about 30 h postradiation. No appreciable amount of preD3 could be detected in the circulation in the early hours after UV-B radiation, suggesting the existence of a process responsible for the specific translocation of vitamin D3 from the skin into the circulation.
Animals↗
[Epidemiology of melioidosis in China].
From 1975 to 1989, a total of 73 strains of P. pseudomallei was isolated from the water samples and the pathological samples of human and domestic animals in 13 counties and cities located different latitude from four provinces Qiong, Yue, Gui and Xiang in China. Serological investigation demonstrated that the geographical distribution of the organism had a significant correlation with the positive rate of antibodies against P. pseudomallei and the native foci of the organisms distributed over the southern subtropical zone and the edge of tropical zone in Qiong, Yue and Gui. In endemic areas, the positive rates of antibodies against P. pseudomallei in human-beings, horses, oxen and pigs are 3.8%-15.2%, 9.1%-18.4%, 6.6%-33.0% and 35% respectively. The investigation results showed the horses and mules infected by the organism would interfere with quarantine of the animals, meanwhile, the meat contaminated by the bacterium would endanger the public health. In Sept. and Oct. of 1989, three cases in Zhanjiang and Sanya of Hainan were reported, two cases died of acute melioidosis with septicemia, another case was the chronic leg ulcers. So, it was predicated that there could have some cases of melioidosis which were misdiagnosed or missed out.
Animals↗
[Glutamyltranspeptidase (gamma-GT) activity in the serum and pharynx tissue of nasopharyngeal carcinoma patients].
The gamma-GT activity in the serum and pharynx tissue of both NPC patients (106) and pharyngitis patients (65) were detected with method of hydrolysis gamma-glutamylnitrophenylamine. The 34 NPC patient's gamma-GT were assayed both before and after radiotherapy. The results indicated that the gamma-GT activities between the NPC patients and pharyngitis patients, and the activities before and after radiotherapy have significant differences. The investigation suggests that the determination of serum and pharynx tissue gamma-GT levels of NPC patients would help to diagnose and evaluate the curative effect for NPC.
Humans↗
[The presence and distribution of insulin in rat brain and its relation to feeding].
Using the immunohistochemical method, insulin was proved to exist in cells of hypothalamus, thalamus, cerebral cortex, hippocampus, ependyma of third ventricle and choroid of rat, including some of the neuronal nuclei, and was further confirmed by immunoelectromicroscopy. The insulin concentrations in rat brain differed in various regions. Simultaneous determinations of insulin concentrations in brain, CSF and blood showed that the brain insulin could not originate from blood through CSF. The participation of brain insulin in regulating of body energy balance at hypothalamus was illustrated and discussed.
Animals↗
[The pressor response induced by electrical stimulation of hypothalamic paraventricular nucleus in the cat].
The experiments were performed on 24 adult cats anesthetized with chloralose and urethane, paralyzed and artificially ventilated. Electrical stimulation of the hypothalamic paraventricular nucleus (PA) with a 12s train of 0.1-0.5 mA and 0.2ms square wave pulses at a frequency of 80 Hz induced a marked increase in arterial blood pressure (n = 19). The 22 points stimulated with maximum responses were clustered in the posterior dorsomedial part of the PA, among them 11 points were associated with an increase in heart rate and 7 a decrease. The remaining 4 points were associated with no change of heart rate, and situated in the anterior part of the PA. Electrolytic lesion of bilateral caudal half of the nucleus of solitary tract (NTS) had no significant effect on cardiovascular responses elicited from electric stimulation of the PA (n = 5). The results suggest that the PA is one of the important centers regulating cardiovascular activity; the NTS may not be involved in the PA pressor response.
Animals↗
[A new natural focus of scrub typhus found in Hunchun].
From May to June 1992, Apodemus agrarius and Apodemus speciosus were captured in Hunchun, Jilin Province. Four strains of Rickettsia tsutsugamushi were isolated from viscera of rats and trombiculid mites. At the same time, the antibody against Rickettsia tsutsugamushi was assayed in the sera of the local people and the wild rats. The positive rates were 15.2% and 16.4%, respectively. The above results showed that a natural focus of scrub typhus exists in Hunchun area.
Animals↗
Rapid method for recovery of DNA from agarose gels.
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Bacteriophage lambda↗
The deduced amino-acid sequence of the cloned cpxR gene suggests the protein is the cognate regulator for the membrane sensor, CpxA, in a two-component signal transduction system of Escherichia coli.
The cpxA gene of Escherichia coli K-12 encodes a membrane-associated sensor element of a two-component signal transduction system in bacteria. The cognate regulator element, however, has not yet been definitively identified. A 2.1-kb segment upstream from cpxA was amplified by polymerase chain reaction, cloned and sequenced. An open reading frame encoding 232 amino acids was found. It showed high homology to the regulator elements of two-component transduction systems. The newly identified gene, designated as cpxR, may encode the cognate protein receiving signals from CpxA.
Amino Acid Sequence↗
Dihydropyrimidine dehydrogenase activity in human peripheral blood mononuclear cells and liver: population characteristics, newly identified deficient patients, and clinical implication in 5-fluorouracil chemotherapy.
Dihydropyrimidine dehydrogenase (DPD) is the initial and rate-limiting enzyme in the catabolism of 5-fluorouracil (FUra), one of the most widely used anticancer drugs. Previous studies from our laboratory demonstrated the clinical importance of DPD in cancer patients (G. D. Heggie, J-P. Sommadossi, D. S. Cross, W. J. Huster, and R. B. Diasio. Cancer Res., 47: 2203-2206, 1987; B. E. Harris, R. Song, S-j. Soong, and R. B. Diasio. Cancer Res., 50: 197-201, 1990), particularly in those with DPD deficiency who experience severe FUra toxicity (including death) following FUra treatment [R. B. Diasio, T. L. Beavers, and J. T. Carpenter. J. Clin. Invest., 81: 47-51, 1988; B. E. Harris, J. T. Carpenter, and R. B. Diasio. Cancer (Phila.), 68: 499-501, 1991]. We now suggest that measurement of DPD activity may be useful in routine screening of cancer patients prior to FUra treatment. In this paper, we describe the following serial studies: (a) we developed a sensitive, accurate, and precise DPD assay and a storage method to stabilize DPD activity, permitting large scale DPD screening in cancer patients; (b) we demonstrated a normal distribution (Gaussian distribution) of human DPD activity from peripheral blood mononuclear cells (PBM-DPD) in a population study. Baselines for PBM-DPD with fresh and frozen samples were 0.425 +/- 0.124 (SD) and 0.189 +/- 0.064 nmol/min/mg protein, respectively. The 95% and 99% distribution ranges for both fresh and frozen samples were also determined, providing criteria for detection of DPD-deficient patients; (c) we identified nine new patients with profound or partial DPD deficiency; (d) we determined a baseline for human liver DPD activity, which was shown to be 0.360 +/- 0.182 nmol/min/mg protein (frozen samples); (e) we did a preliminary evaluation of liver DPD from deficient patients. Low liver DPD activity in two deficient patients correlated with low PBM-DPD activity. Using a polyclonal antibody raised against human liver DPD in our laboratory (Z. Lu, R. Zhang, and R. B. Diasio. J. Biol. Chem., 267: 17102-17109, 1992), Western blot analysis demonstrated decreased DPD protein in the liver cytosol from DPD-deficient patients compared to normal subjects. These results may be useful in improving the effectiveness and/or lessening the toxicity of FUra chemotherapy.
Adult↗
Transcripts for the high and low affinity hexose transporters in rat myoblasts.
The present investigation examined the glucose transporter (GLUT) transcript levels in glucose-grown rat L6 myoblasts, which possessed a high (HAHT) and a low (LAHT) affinity hexose transport system. The most prominent GLUT transcript detected in these cells was a GLUT 3-recognizable 4.1-kilobase transcript. Mutants defective in HAHT contained only residual levels of the GLUT 3 transcript and transporter. Corresponding changes in GLUT 3 transcript levels and HAHT activity were observed in myotubes, confluent cultures, and myogenesis-impaired cells. The other prominent GLUT transcripts detected were the GLUT 4-recognizable 2.8- and 1.4-kilobase transcripts. The close correlation between the GLUT 4 isoform and LAHT activity was indicated by the absence of this isoform in LAHT- mutants, and their corresponding changes in myoblasts grown under different conditions. These findings suggested the HAHT and LAHT transporters might be coded or regulated by the genes responsible for the GLUT 3 and 4 transcripts, respectively. Although relatively high levels of the GLUT 1 transcript and transporter were detected in glucose-grown HAHT-LAHT- mutants, these cells did not exhibit any GLUT 1 transport activity. Since the GLUT 1 transporter was not functional, it was not likely responsible for the HAHT and LAHT activities observed in glucose-grown L6 myoblasts.
Animals↗
An African swine fever virus gene with a similarity to eukaryotic RNA polymerase subunit 6.
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African Swine Fever Virus↗
Relationship between circadian-dependent toxicity of 5-fluorodeoxyuridine and circadian rhythms of pyrimidine enzymes: possible relevance to fluoropyrimidine chemotherapy.
Previous studies in experimental animals and patients have suggested a circadian variation in host toxicity following administration of 5-fluorodeoxyuridine (FdUrd) although the biochemical mechanisms are not fully understood. Thymidine kinase (TK; EC 2.7.1.21), the initial enzyme in the thymidine-phosphorylation pathway, is the first enzyme in the anabolism of FdUrd. Dihydropyrimidine dehydrogenase (DPD; EC 1.3.1.2), is the rate-limiting enzyme in the pyrimidine catabolic pathway and has been shown to be the key enzyme in FdUrd catabolism. The present study examined the relationship between the suggested circadian variation in FdUrd toxicity and potential circadian variations in the activity of these enzymes. Initial studies in Sprague-Dawley rats confirmed that the time of FdUrd administration affected death rate and other drug-related toxicities including loss of body weight, diarrhea, and bone marrow suppression, with the least toxicity and highest survival rate being observed in rats receiving FdUrd at 12:00 noon and 4:00 p.m. and the greatest toxicity and lowest survival rate at 12:00 midnight and 4:00 a.m. Statistical analysis revealed a circadian pattern in FdUrd toxicity (Cosinor analysis, P < 0.001). In subsequent studies with the same species, we simultaneously measured TK and DPD activities in several tissues at various times over 24 h. Under standardized light conditions (lights on, 6:00 a.m. to 6:00 p.m.; lights off, 6:00 p.m. to 6:00 a.m.), with sampling at 4-h intervals (4:00 and 8:00 a.m.; 12:00 noon; 4:00 and 8:00 p.m., and 12:00 midnight), a circadian variation in TK activity was observed (P < 0.0001, Cosinor analysis) in bone marrow, intestinal mucosa, liver, and spleen. In the same group of animals, a circadian pattern of DPD activity in liver and bone marrow was also observed (Cosinor analysis, P < 0.0001) that was inverse compared to the circadian variation in TK activity (Pearson correlation analysis, P < 0.05). Further statistical analysis indicated that the observed circadian variation in FdUrd toxicity was correlated with the circadian variation of TK activity and inversely correlated with DPD activity (Pearson correlation analysis, P < 0.05). Based on the above data, we conclude that the circadian pattern of TK and DPD activity may explain the observed circadian variation in toxicity as the time of FdUrd administration is varied. These results may be useful in the design of improved chemotherapeutic regimens using time-modified administration of FdUrd.
Animals↗
The identification of an African swine fever gene with conserved helicase motifs and a striking homology to herpes virus origin binding protein, UL9.
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African Swine Fever Virus↗
Lysine 87 in the beta subunit of tryptophan synthase that forms an internal aldimine with pyridoxal phosphate serves critical roles in transimination, catalysis, and product release.
This study provides valuable insights into the functions of the lysine residue that forms an internal aldimine with pyridoxal phosphate in the beta subunit of tryptophan synthase from Salmonella typhimurium. Our spectroscopic and kinetic studies demonstrate that a mutant alpha 2 beta 2 complex having beta subunit lysine 87 replaced by threonine forms external aldimines with several amino acids including L-serine, beta-chloro-1-alanine, L-tryptophan, and D-tryptophan. Because the rates of aldimine formation are very slow, we conclude that one role of lysine 87 in the wild type enzyme is to facilitate formation of external aldimines by transimination. Lysine 87 is an essential catalytic residue because the mutant alpha 2 beta 2 complex has no measurable activity in reactions catalyzed by the beta subunit and does not convert external aldimines to products. The mutant enzyme carries out two slow partial beta-elimination reactions: the conversion of beta-chloro-L-alanine and L-serine to enzyme-bound aminoacrylate. The reaction with L-serine is catalyzed by ammonia, which partially replaces the deleted epsilon-amino group. Lysine 87 is important for substrate and product release because L-serine, L-tryptophan, and aminoacrylate dissociate very slowly from the mutant alpha 2 beta 2 complex. Our ability to prepare very stable derivatives of the mutant alpha 2 beta 2 complex containing tightly bound aldimines with a substrate, a product, or a reaction intermediate provides valuable materials for ongoing x-ray crystallographic investigations and future kinetic analyses of the allosteric activation of the alpha subunit by beta subunit ligands.
Catalysis↗
An African swine fever virus gene with similarity to bacterial DNA binding proteins, bacterial integration host factors, and the Bacillus phage SPO1 transcription factor, TF1.
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Amino Acid Sequence↗
Circadian rhythm of rat spleen cytoplasmic thymidine kinase.
The activity of thymidine kinase (TK, EC 2.7.1.21), the first enzyme of the thymidine phosphorylation pathway, was measured at various times over a 24-hr period in the spleens of Sprague-Dawley rats that had been housed under standardized conditions of light and dark for at least 4 weeks before the study. Spleen cytoplasmic TK activity was assayed with [2-14C]thymidine as substrate. Under "normal" light conditions (lights on 6:00 a.m.-6:00 p.m. and lights off 6:00 p.m.-6:00 a.m.), a circadian variation of TK activity was observed (P < 0.0001), Cosinor analysis) with peak activity (1.98 nmol product/hr/mg protein) at 1:00 a.m. (19 hr after light onset, HALO) and trough activity (0.40 nmol product/hr/mg protein) at 1:00 p.m. (7 HALO). Maximum enzyme activity exceeded minimum activity by approximately 5-fold. Reversing the light-dark cycle resulted in a corresponding shift in TK activity. Under these "reverse" conditions (lights on 6:00 p.m.-6:00 a.m. and lights off 6:00 a.m.-6:00 p.m.), a circadian variation in TK activity was also observed (P < 0.0001, Cosinor analysis) with peak activity (1.14 nmol product/hr/mg protein) at 12:00 noon (18 HALO) and trough activity (0.32 nmol/hr/mg protein) at 12:00 a.m. (6 HALO). Maximum enzyme activity exceeded minimum activity by approximately 4-fold. In summary, this study demonstrated for the first time that TK activity varies over a 24-hr period in association with the light-dark cycle.
Animals↗
Investigations of kinase substrate specificity with aqua Rh(III) complexes of adenosine 5'-triphosphate.
In this paper the substrate activities and binding affinities of the stereoisomers of the beta,gamma-bidentate Rh(H2O)4ATP and alpha,beta, gamma-tridentate Rh(H2O)3ATP complexes toward selected members of the kinase family of enzymes are reported. Hexokinase and glycerokinase were found to be specific for the delta beta, gamma-bidentate Rh(H2O)4ATP isomer as substrate while adenylate kinase was found to specifically catalyze the reaction of the delta beta,gamma-bidentate Rh(H2O)4ATP isomer. Pyruvate kinase recognized both the delta beta,gamma-bidentate Rh(H2O)4ATP isomer and the delta beta-P, exo alpha-P alpha,beta,gamma-tridentate Rh(H2O)3ATP isomer as substrates in the catalyzed phosphorylation of the alternate substrate, glycolate. 31P NMR analysis of the respective product complexes showed that alpha-P phosphoryl ligand exchange had not preceded or followed catalysis. Creatine kinase was found to be specific for the delta beta-P, exo alpha-P alpha,beta,gamma-tridentate Rh(H2O)3ATP isomer. Discrimination of the Rh(H2O)nATP isomers via preferential binding of the substrate-active isomer was observed for hexokinase and adenylate kinase but not for glycerokinase, fructose-6 phosphate kinase, creatine kinase, arginine kinase, or acetate kinase.
Adenosine Triphosphate↗