Biomedical subjects

Chuan Li

Publications and source records attributed to Chuan Li.

At least 19 recordsLinked to original sources

PubMed2026

TargetQC: A targeted quality control framework for clinical genomic testing.

Reliable genetic testing depends on accurate assessment of sequencing quality in clinically relevant genomic regions that directly influence variant interpretation. We developed TargetQC, a flexible quality control framework that supports user-defined gene sets, coverage thresholds, and variant sets for evaluating sequencing performance across exome sequencing (ES) and genome sequencing (GS) platforms. TargetQC assesses exon and gene coverage, identifies regions meeting predefined coverage thresholds, evaluates variant detection accuracy, and measures sequencing quality at pathogenic variant sites. We applied TargetQC to the reference sample NA12878 and 665 clinical samples across five ES platforms and one GS platform. ES-VendorB and ES-VendorE achieved the most complete coverage of OMIM coding regions in NA12878, whereas ES-VendorD and ES-VendorE showed the highest coverage compliance in clinical samples. ES-VendorB and GS demonstrated the highest variant detection accuracy. TargetQC provides a practical framework for benchmarking sequencing performance and informing platform selection in clinical genomics.

exome sequencing↗

PubMed2006

The role of N286 and D320 in the reaction mechanism of human dihydrolipoamide dehydrogenase (E3) center domain.

According to the multiple alignment of various dihydrolipoamide dehydrogenases (E3s) sequences, three human mutant E3s of the conserved residues in the center domain, N286D, N286Q, and D320N were created, over-expressed and purified. We characterized these mutants to investigate the reaction mechanism of human dihydrolipoamide dehydrogenases. The specific activities of N286D, N286Q, and D320N are 30.84%, 24.57% and 48.60% to that of the wild-type E3 respectively. The FAD content analysis indicated that these mutant E3s about 96.0%, 99.4% and 82.7% of FAD content compared to that of wild-type E3 respectively. The molecular weight analysis showed that these three mutant proteins form the dimer. Kinetic's data demonstrated that the K(cat) of both forward and reverse reactions of these mutant proteins were decreased. These results suggest that N286 and D320 play a role in the catalytic function of the E3.

Amino Acid Sequence↗

PubMed2006

Synthesis and antitumor evaluation of a novel series of triaminotriazine derivatives.

A series of triaminotriazine derivatives (compounds 5a-f, 6a-x, and 7a-g) was designed, synthesized, and evaluated for their inhibition activities to colorectal cancer (CRC) cell lines (HCT-116 and HT-29). Most of the synthesized compounds demonstrated moderate anti-proliferatory effects on both HCT-116 and HT-29 cell lines at the concentration of 10 microM. The inhibitory activities against HCT-116 and HT-29 cell lines were discussed to develop the structure-activity relationships of this new series. Compounds 6l and 6o exhibited prominent inhibition activities toward HCT-116, with IC50s of 0.76 and 0.92 microM, respectively. The in vivo antitumor studies and pharmacokinetics of compound 6l showed that it might be a promising new hit for further development of antitumor agents.

Antineoplastic Agents↗

PubMed2006

Design and synthesis of novel bis(L-amino acid) ester prodrugs of 9-[2-(phosphonomethoxy)ethyl]adenine (PMEA) with improved anti-HBV activity.

A series of novel bis(L-amino acid) ester prodrugs of 9-[2-(phosphonomethoxy)ethyl] adenine (PMEA) was synthesized and their anti-HBV activity was evaluated in HepG 2 2.2.15 cells. Compounds 11, 12, 21, 22, 26, and 27 demonstrated more potent anti-HBV activity and higher selective index (SI) than adefovir dipivoxil, which was used as a positive control. Compound 11, which was found to be the most potent one, was five times more potent than adefovir dipivoxil with EC50 value of 0.095 microM and CC50 value of 6636 microM. The SI value (>69,000) of compound 11 was 60 times and 24 times higher than those of adefovir dipivoxil and lamivudine, respectively. In vitro stability studies showed that compound 11 was relatively more stable than adefovir dipivoxil with t1/2 of 270 min. These findings suggested that compound 11 could be considered as a promising candidate for further in vivo studies.

Adenine↗

PubMed2006

Characterization and quantification of eight water-soluble constituents in tubers of Pinellia ternata and in tea granules from the Chinese multiherb remedy Xiaochaihu-tang.

In traditional Chinese medicine, multiple herbs are usually used in combination to generate the joint actions of a multiherb remedy. The recent development of LC-hyphenated techniques enables efficient and rapid profiling of the chemical constituent in extracts from multiherb remedies. Xiaochaihu-tang is a seven-herb remedy that has attracted a great deal of attention for reported ability to treat liver dysfunction. Dried tubers of Pinellia ternata (banxia in Chinese) is one of the ingredients, but its chemical contribution to Xiaochaihu-tang remains poorly understood. In the study presented here, LC-UV-MS, LC-MS-MS, and LC-NMR were used in a complementary manner to determine the nature and content of eight water-soluble constituents of banxia and their presence in various tea granules from Xiaochaihu-tang. Among the eight chemicals identified in banxia, cytidine, adenosine, tryptophan, uridine, and adenine are reported for the first time, while tyrosine, guanosine, and phenylalanine were previously described. These chemicals are also present in all of the samples of Xiaochaihu-tang granules, and the amounts of the chemicals ingested due to a daily dose of the multiherb remedies range from 0.008 to 6.3mg.

Adenine↗

PubMed2006

Rapid and direct measurement of free concentrations of highly protein-bound fluoxetine and its metabolite norfluoxetine in plasma.

Fluoxetine (F) and its active N-demethylated metabolite, norfluoxetine (NF), are selective serotonin re-uptake inhibitors that bind extensively to plasma proteins. Development and validation of a novel method for measuring free concentrations of F and NF in plasma are reported here. The plasma filtrate was prepared by a high-speed short-duration ultrafiltration (UF) and then submitted directly to a short-column liquid chromatography/tandem mass spectrometric (LC/MS/MS) assay. There was no significant matrix effect on the analysis, and non-specific binding of the analytes to the UF devices was negligible. For validation of the method, the recovery of the free analytes was compared to that from an optimized equilibrium dialysis method, and analyte stability was examined under conditions mimicking the sample storage, handling, and analysis procedures. The linearity range was 0.37-12 ng/mL for F and NF; the within-run and between-run relative standard deviations were less than 11.9%, and accuracies across the assay range were 100 +/- 10.3%. This new method was then further validated in a pharmacokinetic (PK) study in beagle dogs receiving a single oral dose of fluoxetine hydrochloride. The integrity of the resulting PK data of free F and NF was absolute. The PK data indicate that the novel method is accurate and reliable. To our knowledge this is the first report describing a rapid and reliable method for direct measurement of free concentrations of F and NF in plasma, which will be useful for clinical pharmacokinetic/pharmacodynamic studies of F. Furthermore, the strategies described herein may be applied to the development and validation of methods for measuring the free concentrations of other drugs in plasma.

Animals↗

PubMed2006

The deformation of an erythrocyte under the radiation pressure by optical stretch.

In this paper, the mechanical properties of erythrocytes were studied numerically based upon the mechanical model originally developed by Pamplona and Calladine (ASME J. Biomech. Eng., 115, p. 149, 1993) for liposomes. The case under study is the erythrocyte stretched by a pair of laser beams in opposite directions within buffer solutions. The study aims to elucidate the effect of radiation pressure from the optical laser because up to now little is known about its influence on the cell deformation. Following an earlier study by Guck et al. (Phys. Rev. Lett., 84, p. 5451, 2000; Biophys. J., 81, p. 767, 2001), the empirical results of the radiation pressure were introduced and imposed on the cell surface to simulate the real experimental situation. In addition, an algorithm is specially designed to implement the simulation. For better understanding of the radiation pressure on the cell deformation, a large number of simulations were conducted for different properties of cell membrane. Results are first discussed parametrically and then evaluated by comparing with the experimental data reported by Guck et al. An optimization approach through minimizing the errors between experimental and numerical data is used to determine the optimal values of membrane properties. The results showed that an average shear stiffness around 4.611x10-6 Nm(-1), when the nondimensional ratio of shear modulus to bending modulus ranges from 10 to 300. These values are in a good agreement with those reported in literature.

Cell Size↗

PubMed2006

Maternally derived recombinant human anti-hantavirus monoclonal antibodies are transferred to mouse offspring during lactation and neutralize virus in vitro.

Transgenic mice expressing a recombinant human monoclonal antibody (rHMAb) against hantavirus were generated. These mice could be used as models to explore the possibilities of producing rHMAbs for therapeutic purposes. The highest concentration of the rHMAb in the milk of the transgenic females was 6.6 mg/ml. The rHMAb was also detected in the sera of pups fed by the transgenic females. Both the rHMAbs in the milk of transgenic mice and those in the sera of suckling pups were found to be active against hantaviruses, although the light chain of the antibody absorbed by the pups was modified by N-linked glycosylation.

Animals↗

PubMed2005

S-adenosyl-L-homocysteine hydrolase inactivation curtails ovalbumin-induced immune responses.

The reversible S-adenosyl-L-homocysteine (AdoHcy) hydrolase inhibitor methyl 4-(adenin-9-yl)-2-hydroxybutanoate (DZ2002) suppresses macrophage activation and function. The effects of DZ2002 on T cell function, however, are still unclear. Here, we examined whether DZ2002 alters type 1 helper T cell (Th1) and/or type 2 helper T cell (Th2) immune responses, and whether these effects are associated with both the inhibition of AdoHcy hydrolase and intracellular elevation of endogenous AdoHcy. Male C57BL/6 mice immunized with ovalbumin (OVA) were treated with DZ2002 (1, 5, and 25 mg/kg/day) after which lymphocyte proliferation, cytokine production, and IgG responses to OVA were monitored. Administration of DZ2002 dose dependently suppressed OVA-specific lymphocyte proliferation and anti-OVA IgG production compared with controls. Interleukin (IL)-2 and interferon (IFN)-gamma as well as anti-OVA IgG2a and IgG3, indicators of Th1 immune responses, were markedly decreased in mice treated with DZ2002, whereas IL-4 and anti-OVA IgG1, indicators of Th2 immune responses, were only mildly suppressed. AdoHcy hydrolase activity in spleens of DZ2002-treated mice was substantially blocked, and not surprisingly, AdoHcy levels were significantly elevated compared with controls. Finally, similar immunosuppressive effects were also observed in mice treated with AdoHcy. These data strongly indicate that DZ2002 suppresses antigen-induced specific immune responses, particularly Th1 responses, through inhibition of AdoHcy hydrolase and elevation of endogenous AdoHcy.

Adenine↗

PubMed2005

Dynamic expression of Hsp27 in the presence of mutant ataxin-3.

Machado-Joseph disease (MJD)/spinocerebellar ataxia type 3 (SCA3) is an autosomal dominant spinocerebellar degeneration characterized by a wide range of clinical manifestations. The molecular mechanisms underlying the selective neuronal death typical of MJD/SCA3 are unknown. In this study, human SK-N-SH neuroblastoma cells stably transfected with full-length MJD with 78 CAG repeats were assayed for the dynamic expression of Hsp27, known as a suppressor of poly-Q mediated cell death, in the presence of mutant ataxin-3 in different passages of cultured cells. A dramatic decrease of Hsp27 expression was observed in the earlier passage of cultured SK-N-SH-MJD78 cells, however, the later passage of cells showed a significant increase of Hsp27 to almost the same level of the parental cells. Furthermore, immunohistochemical analysis of MJD transgenic mice and post-mortem human brain tissues showed increased expression of Hsp27 compared to normal control brain, suggesting an up-regulation of Hsp27 in the end stage of MJD. However, mutant cells of earlier passages were more susceptible to serum deprivation than mutant cells of later passages, indicating weak tolerance toward stress in cells with reduced Hsp27. While heat shock was used to assess the stress response, cells expressing mutant ataxin-3 displayed normal response upon heat shock stimuli when compared to the parental cells. Taken together, we proposed that during the early disease stage, the reduction of Hsp27 synthesis mitigated the ability of neuron cells to cope with cytotoxicity induced by mutant ataxin-3, triggering the cell death process during the disease progress. In the late stage of disease, after prolonged stressful conditions of polyglutamine cytotoxicity, the increased level of Hsp27 may reflect a dynamic process of the survived cells to unfold and remove mutant ataxin-3. However, this increased Hsp27 still cannot reverse the global dysfunction of cellular proteins due to accumulation of cytotoxic effects.

Animals↗

PubMed2005

Decreased expression of Hsp27 and Hsp70 in transformed lymphoblastoid cells from patients with spinocerebellar ataxia type 7.

Spinocerebellar ataxia type 7 (SCA7) is caused by an expansion of unstable CAG repeats within the coding region of the novel gene, ataxin-7, on chromosome 3. This disease is also associated with an accumulation of abnormal proteins, including expanded polyglutamine-containing proteins, molecular chaperones, and the ubiquitin-proteasome system. In this study, two SCA7 lymphoblastoid cell lines (LCLs) with 100 and 41 polyglutamine repeats were utilized to examine the effects of polyglutamine expansion on heat shock proteins. Interestingly, under basal conditions, Western blot and immunocytochemical analysis showed a significant decrease of Hsp27 and Hsp70 protein expression in cells containing expanded ataxin-7, as compared with that of the normal LCL. On the other hand, the protein levels of Hsp60 and Hsp90 were not significantly altered in the mutant LCLs. Results from semi-quantitative RT-PCR indicated that the differences in Hsp70 protein levels were due to transcriptional defects while the reduction of Hsp27 in the mutant cells was not caused by transcriptional defects. Our results further demonstrated that despite of defective protein expression of Hsp27 and Hsp70, a normal heat shock response was observed in lymphoblastoid cells expressing mutant ataxin-7. Taken together, our results indicated that expanded ataxin-7 that leads to neurodegeneration significantly impaired the expression of Hsp27 and Hsp70 protein, which may be, at least in part, responsible for the toxicity of mutant ataxin-7 proteins and ultimately resulted in an increase of stress-induced cell death.

Ataxin-7↗

PubMed2005

Vestibular afferent innervation in the vestibular efferent nucleus of rats.

To delineate the vestibular afferent innervation in the vestibular efferent nucleus in the brainstem, neurobiotin or biotinylated dextran amine was injected into the superior Scarpa's ganglion of Sprague-Dawley rats. The locations of vestibular efferent neurons in the brainstem were identified by neutral red or choline acetyltransferase staining. Of the three pairs of vestibular efferent nuclei, labeled fibers and bouton-like endings were found only within the dorsolateral vestibular efferent nucleus on the ipsilateral side. Labeled afferent terminals with bouton-like varicosities were observed in the vicinity of cell bodies or dendrites of these efferent neurons. Our findings suggest that vestibular primary afferents may exert direct influence on vestibular efferent neurons, constituting an ipsilateral close-loop arrangement in the central vestibular system.

Animals↗

PubMed2005

Synthesis and antitumor activity of the hexacyclic camptothecin derivatives.

A series of hexacyclic camptothecin derivatives were synthesized to test for antitumor activity as topoisomerase I inhibitor. The strategy of synthesis was used for the formation of additional furan and dihydrofuran rings fused with 9- and 10-positions of camptothecin. All of the hexacyclic camptothecins were assayed for cytotoxicity against four human tumor cell lines, HL60, BEL-7402, HCT-116, and HeLa, and showed very impressive cytotoxicity activity in vitro. Enzyme activity of the hexacyclic camptothecins was evaluated, being equal or superior to that of SN-38. The stability of four compounds was assessed in human plasma. Two of these compounds were chosen to test for antitumor activity in vivo against Sarcoma-180. The results suggested that additional furan and dihydrofuran rings could improve the antitumor activity in vitro and vivo, though the stability of the lactone ring did not increase.

Animals↗

PubMed2005

Low RET mutation frequency and polymorphism analysis of the RET and EDNRB genes in patients with Hirschsprung disease in Taiwan.

Hirschsprung disease (HSCR), or congenital intestinal aganglionosis, is a relatively common disorder characterized by the absence of ganglion cells in the nerve plexuses of the lower digestive tract, resulting in intestinal obstruction in neonates. Mutations in genes of the RET receptor tyrosine kinase and endothelin receptor B (EDNRB) signaling pathways have been shown to be associated in HSCR patients. In this study, we collected genomic DNA samples from 55 HSCR patients in central Taiwan and analyzed the coding regions of the RET and EDNRB genes by PCR amplification and DNA sequencing. In the 55 patients, an A to G transition was detected in two (identical twin brothers). The mutation was at the end of RET exon 19 at codon 1062 (Y1062C), a reported critical site for the signaling pathways. Single nucleotide polymorphisms (SNP) in exons 2, 7, 11, 13, and 15 of RET and exon 4 of EDNRB in the HSCR patients or controls were detected. The differences between patients and controls in allele distribution of the five RET polymorphic sites were statistically significant. The most frequent genotype encompassing exons 2 and 13 SNPs (the polymorphic sites with the highest percentage of heterozygotes) was AA/GG in patients, which was different from the AG/GT in the normal controls. Transmission disequilibrium was observed in exons 2, 7, and 13, indicating nonrandom association of the susceptibility alleles with the disease in the patients. This study represents the first comprehensive genetic analysis of HSCR disease in Taiwan.

Case-Control Studies↗

PubMed2005

Synthesis and antitumor activity of 7-ethyl-9-alkyl derivatives of camptothecin.

A series of new camptothecin derivatives, as topoisomerase I inhibitor, were synthesized to identify potent antitumor agents. The synthesis method was based on the Claisen rearrangement of 10-allyloxy-7-ethylcamptothecin. All of the compounds were assayed for cytotoxicity against two human tumor cell lines, Bel7402, HCT116, and showed good potency in vitro. Compounds 2, 4, 9, were assessed for the stability of lactone in human plasma. And then compound 2 was tested for antitumor activity in vitro against mouse tumor sarcoma-180. The results suggested that the small alkyl groups in the both 7- and 9-positions of camptothecin could promote liposolubility, antitumor activity in vitro and vivo, though did not bring much increase of the stability of lactone.

Antineoplastic Agents, Phytogenic↗

PubMed2005

A new method for measurement of (-)-sophocarpine, a candidate therapeutic for viral myocarditis, in plasma: application to a toxicokinetic study in beagle dogs.

The naturally occurring alkaloid (-)-sophocarpine (SC) has been developed as a novel anti-coxsackieviral agent for potential treatment of viral myocarditis. However, there is currently no rapid, sensitive method available for ascertaining systemic exposure during the course of SC toxicity studies. The development and full validation of the first rapid and sensitive method, based on liquid chromatography with tandem mass spectrometry (LC/MS/MS), for determination of SC in plasma is reported here. This new assay increases sample throughput by using minimal sample clean-up procedures and short chromatographic analysis times. The bio-matrix effect on the ionization of SC and the internal standard, (-)-stepholidine, was investigated for method development and validation, and two organic solvents (methyl tert-butyl ether and ethyl acetate) were found for sample preparation that led to low matrix effects. The new analytical method was used to analyze plasma samples obtained from a repeated-dose toxicity study of SC in beagle dogs. The results of the toxicokinetic analysis indicated that the systemic exposure to SC was proportional to the dose, and that no significant accumulation of SC was observed after 3 months of repeated treatments with intravenous SC at 7.5, 15, or 30 mg/kg/day. This sensitive and specific LC/MS/MS technique can form the basis for accurate quantification of SC in various biological fluids for preclinical and clinical pharmacokinetic evaluation.

Alkaloids↗

PubMed2005

SARS patients-derived human recombinant antibodies to S and M proteins efficiently neutralize SARS-coronavirus infectivity.

OBJECTIVE: To develop a specific SARS virus-targeted antibody preparation for emergent prophylaxis and treatment of SARS virus infection. METHODS: By using phage display technology, we constructed a naive antibody library from convalescent SARS patient lymphocytes. To obtain the neutralizing antibody to SARS virus surface proteins, the library panning procedure was performed on purified SARS virions and the specific Fab antibody clones were enriched by four rounds of repeated panning procedure and screened by highthroughput selection. The selected Fab antibodies expressed in the periplasma of E. coli were soluble and further purified and tested for their binding properties and antiviral function to SARS virus. The functional Fab antibodies were converted to full human IgG antibodies with recombinant baculovirus/insect cell systems and their neutralizing activities were further determined. RESULTS: After four rounds of the panning, a number of SARS-CoV virus-targeted human recombinant Fab antibodies were isolated from the SARS patient antibody library. Most of these were identified to recognize both natural and recombinant SARS spike (S) proteins, two Fab antibodies were specific for the virus membrane (M) protein, only one bound to SARS-CoV nucleocapsid protein. The SARS-CoV S and M protein-targeted Fab or IgG antibodies showed significant neutralizing activities in cytopathic effect (CPE) inhibition neutralization test, these antibodies were able to completely neutralize the SARS virus and protect the Vero cells from CPE after virus infection. However, the N protein-targeted Fab or IgG antibodies failed to neutralize the virus. In addition, the SARS N protein-targeted human Fab antibody reacted with the denatured N proteins, whereas none of the S and M protein specific neutralizing antibodies did. These results suggested that the S and M protein-specific neutralizing antibodies could recognize conformational epitopes which might be involved in the binding of virions to cellular receptors and the fusion activity of the virus. CONCLUSION: The SARS-CoV spike protein and membrane proteins are able to elicite efficient neutralizing antibodies in SARS patients. The neutralizing antibodies we generated in this study may be more promising candidates for prophylaxis and treatment of SARS infection.

Amino Acid Sequence↗

PubMed2004

Identification and phylogenetic analyses of the protein arginine methyltransferase gene family in fish and ascidians.

Protein arginine methyltransferases (PRMT) involved in the regulations of signal transduction, protein subcellular localization, and transcription have been mostly studied in mammals and yeast. In this study orthologues of eight human PRMT genes (PRMT1-7 and HRMT1L3) were identified in both puffer fish Fugu rubripes and zebrafish Danio rerio. The fish PRMT genes appear to be conserved with their mammalian orthologues at the levels of amino acid sequences as well as genomic structures. All vertebrate PRMT genes contain 10-16 coding exons except PRMT6 that contains only one coding exon. Western blot analyses of zebrafish tissue extracts confirmed the expression of some PRMT proteins in zebra fish. We further identified six PRMT members (PRMT1, 3-7) in an invertebrate chordate Ciona intestinalis. Genomic structures of the PRMT orthologues are no more conserved in the ascidians, as PRMT3 and PRMT5 contain only one coding exon while PRMT6 contains six exons. PRMT2 and HRMT1L3 that are missing in Ciona appear to be vertebrate-specific. HRMT1L3 is a PRMT1 paralogue with highly conserved sequences and exact exon junctions, whereas the PRMT2 orthologues are very diverged. Different PRMT orthologues are likely to evolve at different rates and the PRMT1 orthologues appear to be most conserved through evolution. Furthermore, phylogenetic analyses using the core regions of various PRMT genes show that PRMT5 with the type II PRMT activity is separated in one branch. All other PRMT genes including PRMT1, 2, 3, 4, 6, 7 and HRMT1L3 clustered in the other branch, probably represent the genes for the type I activity.

Amino Acid Sequence↗