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Biomedical subjects

C Mao

Publications and source records attributed to C Mao.

At least 55 records · Page 3Linked to original sources
PubMed2000

Structure-based design of a novel synthetic spiroketal pyran as a pharmacophore for the marine natural product spongistatin 1.

SPIKET-P, a novel synthetic spiroketal pyran, was rationally designed as a pharmocophore for the tubulin depolymerizing marine natural product Spongistatin 1. SPIKET-P was prepared from the commercially available benzyl (R)-(-)-glycidyl ether using a versatile 11-step synthetic scheme in a stereocontrolled fashion. At nanomolar concentrations, SPIKET-P caused tubulin depolymerization in cell-free turbidity assays and exhibited potent cytotoxic activity against cancer cells as evidenced by destruction of microtubule organization, and prevention of mitotic spindle formation in human breast cancer cells.

Animals↗
PubMed2000

Cloning of an alkaline ceramidase from Saccharomyces cerevisiae. An enzyme with reverse (CoA-independent) ceramide synthase activity.

Ceramide is not only a core intermediate of sphingolipids but also an important modulator of many cellular events including apoptosis, cell cycle arrest, senescence, differentiation, and stress responses. Its turnover may be tightly regulated. However, little is known about the regulation of its metabolism because most enzymes responsible for its synthesis and breakdown have yet to be cloned. Here we report the cloning and characterization of the yeast gene YPC1 (YBR183w) by screening Saccharomyces cerevisiae genes whose overexpression bestows resistance to fumonisin B1. We demonstrate that the yeast gene YPC1 encodes an alkaline ceramidase activity responsible for the breakdown of dihydroceramide and phytoceramide but not unsaturated ceramide. YPC1 ceramidase activity was confirmed by in vitro studies using an Escherichia coli expression system. Importantly, YPC1p also has reverse activity, catalyzing synthesis of phytoceramide from palmitic acid and phytosphingosine. This ceramide synthase activity is CoA-independent and is resistant to fumonisin B1, thus explaining why YPC1 was cloned as a fumonisin B1-resistant gene.

Amidohydrolases↗
PubMed2000

Cre-mediated cerebellum- and hippocampus-restricted gene mutation in mouse brain.

Using the phage P1-derived Cre/loxP recombination system, we have created a line of cre-transgenic mice in which the Cre-mediated gene deletion is restricted to granule cells of cerebellum and dentate gyrus of hippocampus. Low levels of deletion were also present in pyramidal cells of hippocampal CA1 and CA3 fields. The Cre/loxP recombination occurred prenatally. The recombination efficiencies in the granular layer of the cerebellum, the granular layer of the dentate gyrus, and the CA1 and CA3 pyramidal cells of the hippocampus were 34.0%, 23.1%, 3.0%, and 9.8%, respectively. This line of cre-transgenic mice should be conducive to studies of the effect of a gene mutation upon brain development and plasticity.

Animals↗
PubMed2000

Proteinase inhibitor 9, an inhibitor of granzyme B-mediated apoptosis, is a primary estrogen-inducible gene in human liver cells.

Although liver is an estrogen target tissue, the number of hepatic genes known to be directly induced by estrogen is very small. We identified proteinase inhibitor 9, or PI-9, as being rapidly and strongly induced by estrogen in an estrogen receptor-positive human liver cell line (HepG2-ER7). Since PI-9 mRNA was also induced by estrogen in a human liver biopsy sample, PI-9 is a genuine estrogen-regulated human gene. PI-9 is a potent inhibitor of granzyme B and of granzyme B-mediated apoptosis. Estrogens induced PI-9 mRNA within 2 h, PI-9 mRNA levels reached a plateau of 30-40-fold induction in 4 h, and induction was not blocked by cycloheximide, indicating that induction of PI-9 mRNA is a primary response. The antiestrogen trans-hydroxytamoxifen was a partial agonist for PI-9 mRNA induction, whereas the antiestrogen ICI 182, 780 was a pure antagonist. Western blot analysis showed that estrogen strongly increases PI-9 protein levels. Inhibition of transcription with actinomycin D resulted in identical rates of PI-9 mRNA decay in the presence and absence of estrogen. We isolated genomic clones containing the PI-9 promoter region, identified a putative transcription start site, and carried out transient transfections of PI-9-luciferase reporter gene constructs. The estrogen, moxestrol, elicited a robust induction from the PI-9-luciferase reporter. Mutational inactivation of three potential imperfect estrogen response elements in the PI-9 5'-flanking region had no effect on moxestrol estrogen receptor induction.

Apoptosis↗
PubMed2000

Multimerization-cyclization of DNA fragments as a method of conformational analysis.

Ligation of short DNA fragments results in the formation of linear and circular multimers of various lengths. The distribution of products in such a reaction is often used to evaluate fragment bending caused by specific chemical modification, by bound ligands or by the presence of irregular structural elements. We have developed a more rigorous quantitative approach to the analysis of such experimental data based on determination of j-factors for different multimers from the distribution of the reaction products. j-Factors define the effective concentration of one end of a linear chain in the vicinity of the other end. To extract j-factors we assumed that kinetics of the reaction is described by a system of differential equations where j-factors appear as coefficients. The assumption was confirmed by comparison with experimental data obtained here for DNA fragments containing A-tracts. At the second step of the analysis j-factors are used to determine conformational parameters of DNA fragments: the equilibrium bend angle, the bending rigidity of the fragment axis, and the total twist of the fragments. This procedure is based on empirical equations that connect the conformational parameters with the set of j-factors. To obtain the equations, we computed j-factors for a large array of conformational parameters that describe model fragments. The approach was tested on both simulated and actual experimental data for DNA fragments containing A-tracts. A-tract DNA bend angle determined here is in good agreement with previously published data. We have established a set of experimental conditions necessary for the data analysis to be successful.

Base Sequence↗
PubMed2000

Yeast sphingosine-1-phosphate phosphatases: assay, expression, deletion, purification, and cellular localization by GFP tagging.

DHS-1-P phosphatases cloned from yeast represent novel lipid phosphatases, which were not thought to exist in yeast. Identification and characterization of YSR2 and YSR3 have demonstrated that the DHS-1-P phosphatase is an important mediator in the biosynthesis of sphingolipids and in the maintenance of the balance of signaling lipid molecules ceramide, sphingosine, and sphingosine-1-P. Methods introduced here for purification, activity assay, in vivo labeling, and cellular localization using GFP tagging are expected to facilitate our understanding of this enzyme.

Cell Compartmentation↗
PubMed2000

Differential expression of rat frizzled-related frzb-1 and frizzled receptor fz1 and fz2 genes in the rat aorta after balloon injury.

Frzb-1 is a secreted protein, presenting similarity with the Wnt-binding domain of the frizzled family of receptors, which acts as an antagonist of Wnt signaling. Using mRNA differential display in the rat aorta balloon injury model, we identified overexpression of Frzb-1 mRNA and determined its cDNA sequence. By quantitative reverse transcription-polymerase chain reaction and RNase protection assay, a biphasic upregulation of rFrzb-1 expression was observed, with significant peaks of a 1.7-fold increase at 4 days and a 1. 5-fold increase at 3 weeks after aortic injury in vivo. In contrast, expression of the rat frizzled receptor genes rfz1 and rfz2 were transiently downregulated at 1 and 4 hours after balloon injury. rFrzb-1 was expressed predominantly in rat aortic smooth muscle cells (RASMCs) and barely in aortic fibroblasts and endothelial cells (RAECs), whereas rfz1 and rfz2 were expressed in all of these cells when stimulated with serum. Transient downregulation of rfz1 and rfz2 expression was reproduced by stimulation of quiescent RASMCs with serum, platelet-derived growth factor-BB, or fibroblast growth factor-2. In contrast, rFrzb-1 expression diminished slowly, to reach a 2-fold decrease 24 hours after growth factor stimulation, implying that quiescent RASMCs expressed higher levels of rFrzb-1 mRNA than did proliferative ones. Overexpression of rFrzb-1 in the aorta seemed to coincide with the arrest of RASMC proliferation occurring in the media 4 days and in the neointima 3 weeks after balloon injury. Our results demonstrate that rfrzb-1, rfz1, and rfz2 are differentially regulated in response to arterial injury and that this modulation seems to follow the proliferative state of RASMCs, suggesting that these Wnt-signaling components may be involved in intimal vascular disease.

Animals↗
PubMed2000

N-[2-(4-methylphenyl)ethyl]-N'-[2-(5-bromopyridyl)]-thiourea as a potent inhibitor of NNRTI-resistant and multidrug-resistant human immunodeficiency virus type 1.

The composite non-nucleoside reverse transcriptase inhibitor (NNRTI) binding pocket model was used to study a number of thiourea analogues with different substitutions at the 4-phenyl position including N-[2-(4-methylphenyl)ethyl]-N'-[2-(5-bromopyridyl)]-thiourea (compound HI-244), which inhibited recombinant RT better than trovirdine or compound HI-275 with an unsubstituted phenyl ring. HI-244 effectively inhibited the replication of HIV-1 strain HTLV(IIIB) in human peripheral blood mononuclear cells with an IC50 value of 0.007 microM, which is equal to the IC50 value of trovirdine. Notably, HI-244 was 20 times more effective than trovirdine against the multidrug-resistant HIV-1 strain RT-MDR with a V106A mutation (as well as additional mutations involving the RT residues 74V, 41L and 215Y) and seven times more potent than trovirdine against the NNRTI-resistant HIV-1 strain A17 with a Y181C mutation.

Anti-HIV Agents↗
PubMed2000

Piperidinylethyl, phenoxyethyl and fluoroethyl bromopyridyl thiourea compounds with potent anti-HIV activity.

Derivatives of piperidinylethyl, phenoxyethyl and fluoroethyl bromopyridyl thioureas were designed and synthesized as non-nucleoside reverse transcriptase inhibitors (NNRTIs) of HIV-1 reverse transcriptase (RT). The anti-HIV activity of these compounds was examined by determining their ability to inhibit the replication of the HIV-1 strain HTLV(IIIB) in human peripheral blood mononuclear cells. The unsubstituted parent pyridyl thiourea compound N-[2-(1-piperidine)ethyl]-N'-[2-(pyridyl)] thiourea (1) exhibited no anti-HIV activity, even at 100 microM. However, the thiourea derivatives that contain a bromo- or chloro-substituted pyridyl group, compounds 2 and 5, inhibited HIV-1 replication at nanomolar concentrations. The addition of a methyl group onto the piperidine ring significantly altered the potency of these compounds; while methyl substitution at the 3-position of the piperidine ring reduced the activity, methyl substitution at the 2-position enhanced the anti-HIV activity. The IC50 value of the lead piperidinyl compound, N-[2-(2-methylpiperidinylethyl)]-N'-[2-(5-bromopyridyl)] thiourea (4) was <0.001 microM. All three phenoxyethyl derivatives, including the unsubstituted parent phenoxyethyl pyridyl thiourea compound N-[2-(phenoxy)ethyl]-N'-[2-(pyridyl)]thiourea (8) and the bromo-/chloro-substituted phenoxyethyl halopyridyl thiourea compounds N-[2-(phenoxy)ethyl]-N'-[2-(5-chloropyridyl)]thiourea (9) and N-[2-(phenoxy)ethyl]-N'-[2-(5-bromopyridyl)]thiourea (10) exhibited potent anti-HIV activity with nanomolar IC values. The corresponding fluoroethyl halopyridyl thiourea compounds beta-fluoro[2-phenethyl]-N'[2-(5-chloropyridyl)]thiourea (11) and beta-fluoro[2-phenethyl]-N'[2-(5-bromopyridyl)]thiourea (12) inhibited HIV-1 replication in PBMC with subnanomolar IC50 values and selectivity indices >30000. Compared to the corresponding phenoxyethyl thiourea compounds 9 and 10, these compounds were >4-5-fold more active as anti-HIV agents. Notably, the lead fluorothiourea compounds 11 and 12 were both substantially more active against the NNRTI-resistant HIV strains RT-MDR (V106A) and A17 (Y181C) than nevirapine or delavirdine. Taken together, our results provide additional experimental evidence that the structural features of the 'linker unit' between the pyridyl and phenyl moieties and changes in the phenyl group of PETT-related thiourea compounds significantly affects their biological activity as NNRTIs of HIV-1 RT.

Anti-HIV Agents↗
PubMed2000

A histone deacetylase inhibitor potentiates estrogen receptor activation of a stably integrated vitellogenin promoter in HepG2 cells.

To compare the role of histone deactylation in estrogen activation of a transiently transfected vitellogenin (VIT) promoter and an integrated VIT promoter in the same cells, we produced three HepG2, human hepatoma, cell lines (HepG2ERV cells) stably expressing human estrogen receptor alpha (hERalpha) and containing an integrated VIT promoter-chloramphenicol acetyltransferase (VIT-CAT) reporter gene. The three ER-positive HepG2ERV cell lines and wild-type, ER-negative, HepG2 cells cotransfected with cytomegalovirus-hERalpha exhibited similar MOX-dependent inductions of 20- to 50-fold with a transiently transfected VIT-luciferase reporter and 15- to 50-fold with a transfected 4-estrogen response element-TATA-luciferase reporter gene. The histone deacetylase inhibitor, trichostatin A, did not enhance MOX induction of the transiently transfected VIT promoter in the HepG2ERV cells. In contrast, trichostatin A dramatically potentiated MOX induction of the stably integrated VIT-CAT reporter gene, resulting in MOX-ER-dependent increases in CAT activity of up to 600-fold. These data demonstrate that although liganded ER exhibits the capacity to fully activate a transiently transfected VIT promoter, under some circumstances the ability to reorganize a repressive chromatin structure may be limiting for steroid receptor action.

Chloramphenicol O-Acetyltransferase↗
PubMed2000

SPIKET and COBRA compounds as novel tubulin modulators with potent anticancer activity.

Agents that either promote or inhibit tubulin polymerization exhibit anticancer activity by disrupting normal mitotic spindle assembly and cell division as well as inducing apoptosis. Recently identified novel agents that target tubulin include synthetic spiroketal pyrans (SPIKET), targeting the spongistatin binding site of beta-tubulin, and COBRA compounds, targeting a unique binding cavity on alpha-tubulin. At nanomolar concentrations, the SPIKET compound SPIKET-P caused tubulin depolymerization in cell-free turbidity assays and exhibited potent cytotoxic activity against cancer cells as evidenced by destruction of microtubule organization, and prevention of mitotic spindle formation in human breast cancer cells. Molecular modeling studies predicted a high-affinity interaction of the first COBRA compounds, COBRA-0 and COBRA-1, with a unique hydrophobic binding site on alpha-tubulin located between the GTP/GDP binding site and the M-loop. Further studies showed that COBRA-1 inhibited GTP-induced tubulin polymerization in cell-free tubulin turbidity assays. Treatment of human breast cancer and brain tumor (glioblastoma) cells with COBRA-1 caused destruction of microtubule organization and apoptosis. COBRA-1 activated the pro-apoptotic c-Jun N-terminal kinase (JNK) signal transduction pathway. COBRA and SPIKET compounds represent two new classes of tubulin targeting agents that show promise as anticancer drugs.

Animals↗
PubMed2000

[Simultaneous primary palate repair and alveolar bone grafting in unoperated cleft palate patients over 8 years old].

OBJECTIVE: To analyze the applicability and results of simultaneous primary palate repair and alveolar bone grafting in unoperated cleft lip and palate patients over 8 years old. METHODS: A retrospective study was performed in a group of unoperated cleft palate patients who received simultaneous primary palate repair and alveolar bone grafting. Between December 1990 and March 1998, a consecutive of 38 complete unilateral cleft lip and palate patients were treated by the procedures of simultaneous primary cleft palate repair and alveolar bone grafting at the Peking University of Cleft Lip and Palate Treatment Center. All the patients had their lip repaired before they were admitted. The age range was 8 to 24 years, with the average of 14.7 years old. The duration of operation as well as the blood loss during the operation was recorded, and compared with those patients who only received alveolar bone grafting. All the patients have been followed up for at least twelve months, and the results of bone grafting were evaluated according to radiographs. RESULTS: All the operations were successful, and the wound healed well. Compared with simple alveolar bone grafting, simultaneous primary palate repair and alveolar bone grafting prolonged the operation time to an average of 37 minutes. The procedure of simultaneous primary palate repair and alveolar bone grafting did not prolong the operating time compared with simple alveolar bone grafting and no blood transfusion due to bone grafting was needed. All the wounds both in the grafted area and donor site healed uneventfully. No major complications occurred during or shortly after the operation. The overall clinical successful rate of alveolar bone grafting in this group of patients was 89.5%. CONCLUSION: Simultaneous primary palate repair and alveolar bone grafting are safe and applicable procedure for unoperated cleft palate patients, and this procedure should be performed in those unoperated cleft palate patients above 8 years old.

Adolescent↗
PubMed2000

[Studies on antiplatelet aggregation and anticoagulant action of Curcuma phaeocaulis].

OBJECTIVE: To study the effects of Curcuma phaeocaulis Valeton on platelet aggregation and anticoagulant action. METHODS: Antiplatelet aggregation and anticoagulant action of Curcuma phaeocaulis Valeton of mice is observed in platelet aggregation and coagulant time. RESULTS: The experimental results show that Curcuma phaeocaulis Valeton can inhibit platelet aggregation, prolong time of mice coagulation. CONCLUSION: Antiplatelet aggregation and anticoagulant action of processed products with vinegar is the most powerful.

Animals↗
PubMed2000

A retrospective study of bilateral alveolar bone grafting.

OBJECTIVE: To evaluate the treatment results of bilateral alveolar bone grafting (BABG) in patients with bilateral complete clefts of lip and palate. METHODS: A retrospective study was performed in 66 bilateral complete cleft lip and palate patients who received the procedure of BABG, among them 15 were primary BABG and 51 were secondary BABG. The patients were further divided into three groups according to age and eruption stage of the canine at the time of surgery. The result of BABG was evaluated on the radiographs. RESULTS: (1) The overall success rate of BABG was 75.0%, with 83.3% and 72.5% for primary and secondary BABG respectively; (2) The marginal bone level was found to be significantly higher in the youngest age group than in the other groups both for primary and secondary BABG; (3) For both primary and secondary BABG, Group C (patients' age more than 16 years) had the least optimal success rate, with 66.7% and 65.1% respectively. CONCLUSION: Simultaneous primary palate repair and BABG is safe and feasible procedure for treating unoperated bilateral complete cleft lip and cleft palate patients. For both primary and secondary BABG, significantly better results can be achieved if the operation is performed before eruption of the canine.

Adolescent↗
PubMed1999

N'-[2-(2-thiophene)ethyl]-N'-[2-(5-bromopyridyl)] thiourea as a potent inhibitor of NNI-resistant and multidrug-resistant human immunodeficiency virus-1.

The thiophene-ethyl thiourea (TET) compound N'-[2-(2-thiophene)ethyl]-N'-[2-(5-bromopyridyl)]-thiourea (compound HI-443) was five times more potent than trovirdine, 1250 times more potent than nevirapine, 100 times more potent than delavirdine, 75 times more potent than MKC-442, and 50 times more potent than AZT against the multidrug resistant HIV-1 strain RT-MDR with a V106A mutation. HI-443 was almost as potent against the NNI-resistant HIV-1 strain A17 with a Y181C mutation as it was against HTLV(IIIB). The activity of HI-443 against A17 was ten times more potent than that of trovirdine, 2083 times more potent than that of nevirapine, and 1042 times more potent than that of delavirdine. HI-443 inhibited the replication of the NNI-resistant HIV-1 strain A17 variant with Y181C plus K103N mutations in RT with an IC50 value of 3.3 microM, whereas the IC50 values of trovirdine, nevirapine, and delavirdine were all >100 microM. These findings establish the novel thiophene containing thiourea compound HI-443 as a novel NNI with potent antiviral activity against NNI-sensitive, NNI-resistant and multidrug-resistant strains of HIV-1.

Anti-HIV Agents↗
PubMed1999

No braiding of Holliday junctions in positively supercoiled DNA molecules.

The Holliday junction is a prominent intermediate in genetic recombination that consists of four double helical arms of DNA flanking a branch point. Under many conditions, the Holliday junction arranges its arms into two stacked domains that can be oriented so that genetic markers are parallel or antiparallel. In this arrangement, two strands retain a helical conformation, and the other two strands effect the crossover between helical domains. The products of recombination are altered by a crossover isomerization event, which switches the strands fulfilling these two roles. It appears that effecting this switch from the parallel conformation by the simplest mechanism results in braiding the crossover strands at the branch point. In previous work we showed by topological means that a short, parallel, DNA double crossover molecule with closed ends did not braid its branch point; however, that molecule was too short to adopt the necessary positively supercoiled topology. Here, we have addressed the same problem using a larger molecule of the same type. We have constructed a parallel DNA double crossover molecule with closed ends, containing 14 double helical turns in each helix between its crossover points. We have prepared this molecule in a relaxed form by simple ligation and in a positively supercoiled form by ligation in the presence of netropsin. The positively supercoiled molecule is of the right topology to accommodate braiding. We have compared the relaxed and supercoiled versions for their responses to probes that include hydroxyl radicals, KMnO4, the junction resolvases endonuclease VII and RuvC, and RuvC activation of KMNO4 sensitivity. In no case did we find evidence for a braid at the crossover point. We conclude that Holliday junctions do not braid at their branch points, and that the topological problem created by crossover isomerization in the parallel conformation is likely to be solved by distributing the stress over the helices that flank the branch point.

Bacterial Proteins↗
PubMed1999

N-[2-(1-cyclohexenyl)ethyl]-N'-[2-(5-bromopyridyl)]-thiourea and N'-[2-(1-cyclohexenyl)ethyl]-N'-[2-(5-chloropyridyl)]-thiourea as potent inhibitors of multidrug-resistant human immunodeficiency virus-1.

We have replaced the pyridyl ring of trovirdine with an alicyclic cyclohexenyl, adamantyl or cis-myrtanyl ring. Only the cyclohexenyl-containing thiourea compound N-[2-(1-cyclohexenyl)ethyl]-N'-[2-(5-bromopyridyl)]- thiourea (HI-346) (as well as its chlorine-substituted derivative N-[2-(1-cyclohexenyl)ethyl]-N'-[2-(5-chloropyridyl)]- thiourea/HI-445) showed RT inhibitory activity. HI-346 and HI-445 effectively inhibited recombinant RT with better IC50 values than other anti-HIV agents tested. The ranking order of efficacy in cell-free RT inhibition assays was: HI-346 (IC50 = 0.4 microM) > HI-445 (IC50 = 0.5 microM) > trovirdine (IC50 = 0.8 microM) > MKC-442 (IC5 = 0.8 microM) = delavirdine (IC50 = 1.5 microM) > nevirapine (IC50 = 23 microM). In accord with this data, both compounds inhibited the replication of the drug-sensitive HIV-1 strain HTLV(IIIB) with better IC50 values than other anti-HIV agents tested. The ranking order of efficacy in cellular HIV-1 inhibition assays was: HI-445 = HI-346 (IC50 = 3 nM) > MKC-442 (IC50 = 4 nM) = AZT (IC50 = 4 nM) > trovirdine (IC50 = 7 nM) > delavirdine (IC50 = 9 nM) > nevirapine (IC50 = 34 nM). Surprisingly, the lead compounds HI-346 and HI-445 were 3-times more effective against the multidrug resistant HIV-1 strain RT-MDR with a V106A mutation (as well as additional mutations involving the RT residues 74V,41L, and 215Y) than they were against HTLV(IIIB) with wild-type RT. HI-346 and HI-445 were 20-times more potent than trovirdine, 200-times more potent than AZT, 300-times more potent than MKC-442, 400-times more potent than delavirdine, and 5000-times more potent than nevirapine against the multidrug resistant HIV-1 strain RT-MDR. HI-445 was also tested against the RT Y181C mutant A17 strain of HIV-1 and found to be >7-fold more effective than trovirdine and >1,400-fold more effective than nevirapine or delavirdine. Similarly, both HI-346 and HI-445 were more effective than trovirdine, nevirapine, and delavirdine against the problematic NNI-resistant HIV-1 strain A17-variant with both Y181C and K103N mutations in RT, although their activity was markedly reduced against this strain. Neither compound exhibited significant cytotoxicity at effective concentrations (CC50 >100 microM). These findings establish the lead compounds HI-346 and HI-445 as potent inhibitors of drug-sensitive as well as multidrug-resistant stains of HIV-1.

Aminopyridines↗