Biomedical subjects

Lili Chen

Publications and source records attributed to Lili Chen.

33 records · Page 2Linked to original sources

PubMed2005

Cinanserin is an inhibitor of the 3C-like proteinase of severe acute respiratory syndrome coronavirus and strongly reduces virus replication in vitro.

The 3C-like proteinase (3CLpro) of severe acute respiratory syndrome-associated coronavirus (SARS-CoV) is one of the most promising targets for anti-SARS-CoV drugs due to its crucial role in the viral life cycle. In this study, a database containing structural information of more than 8,000 existing drugs was virtually screened by a docking approach to identify potential binding molecules of SARS-CoV 3CLpro. As a target for screening, both a homology model and the crystallographic structure of the binding pocket of the enzyme were used. Cinanserin (SQ 10,643), a well-characterized serotonin antagonist that has undergone preliminary clinical testing in humans in the 1960s, showed a high score in the screening and was chosen for further experimental evaluation. Binding of both cinanserin and its hydrochloride to bacterially expressed 3CLpro of SARS-CoV and the related human coronavirus 229E (HCoV-229E) was demonstrated by surface plasmon resonance technology. The catalytic activity of both enzymes was inhibited with 50% inhibitory concentration (IC50) values of 5 microM, as tested with a fluorogenic substrate. The antiviral activity of cinanserin was further evaluated in tissue culture assays, namely, a replicon system based on HCoV-229E and quantitative test assays with infectious SARS-CoV and HCoV-229E. All assays revealed a strong inhibition of coronavirus replication at nontoxic drug concentrations. The level of virus RNA and infectious particles was reduced by up to 4 log units, with IC50 values ranging from 19 to 34 microM. These findings demonstrate that the old drug cinanserin is an inhibitor of SARS-CoV replication, acting most likely via inhibition of the 3CL proteinase.

Animals↗

PubMed2005

Delivery of systemic chemotherapeutic agent to tumors by using focused ultrasound: study in a murine model.

PURPOSE: To quantitatively determine the delivery of systemic liposomal doxorubicin to tumors treated with pulsed high-intensity focused ultrasound and to study the mechanism underlying this delivery in a murine model. MATERIALS AND METHODS: All animal work was performed in compliance with guidelines and approval of institutional animal care committee. C3H mice received subcutaneous injections in the flank of a cell suspension of SCC7, a murine squamous cell carcinoma cell line; mice (n = 32) in drug delivery study received unilateral injections, whereas mice (n = 10) in mechanistic study received bilateral injections. Tumors were treated when they reached 1 cm(3) in volume. In the drug delivery study, doxorubicin hydrochloride liposomes were injected into the tail vein: Mice received therapy with doxorubicin injections and high-intensity focused ultrasound, doxorubicin injections alone, or neither form of therapy (controls). Tumors were removed, and the doxorubicin content was assayed with fluorescent spectrophotometry. In the mechanistic study, all mice received an injection of 500-kDa dextran-fluorescein isothyocyanate into the tail vein, and half of them were exposed to high-intensity focused ultrasound prior to injection. Contralateral tumors served as controls for each group. Extravasation of dextran-fluorescein isothyocyanate was observed by using in vivo confocal microscopy. RESULTS: Mean doxorubicin concentration in tumors treated with pulsed high-intensity focused ultrasound was 9.4 microg . g(-1) +/- 2.1 (standard deviation), and it was significantly higher (124% [9.4 microg . g(-1)/4.2 microg . g(-1)]) than in those that were not treated with high-intensity focused ultrasound (4.2 microg . g(-1) +/- 0.95) (P < .001, unpaired two-tailed Student t test). Extravasation of dextran-fluorescein isothyocyanate was observed in the vasculature of tumors treated with high-intensity focused ultrasound but not in that of untreated tumors. CONCLUSION: Pulsed high-intensity focused ultrasound is an effective method of targeting systemic drug delivery to tumor tissue. Potential mechanisms for producing the observed enhancement are discussed.

Animals↗

PubMed2004

Severe acute respiratory syndrome coronavirus 3C-like proteinase N terminus is indispensable for proteolytic activity but not for enzyme dimerization. Biochemical and thermodynamic investigation in conjunction with molecular dynamics simulations.

Severe acute respiratory syndrome (SARS) coronavirus is a novel human coronavirus and is responsible for SARS infection. SARS coronavirus 3C-like proteinase (SARS 3CL(pro)) plays key roles in viral replication and transcription and is an attractive target for anti-SARS drug discovery. In this report, we quantitatively characterized the dimerization features of the full-length and N-terminal residues 1-7 deleted SARS 3CL(pro)s by using glutaraldehyde cross-linking SDS-PAGE, size-exclusion chromatography, and isothermal titration calorimeter techniques. Glutaraldehyde cross-linking SDS-PAGE and size-exclusion chromatography results show that, similar to the full-length SARS 3CL(pro), the N-terminal deleted SARS 3CL(pro) still remains a dimer/monomer mixture within a wide range of protein concentrations. Isothermal titration calorimeter determinations indicate that the equilibrium dissociation constant (K(d)) of the N-terminal deleted proteinase dimer (262 microm) is very similar to that of the full-length proteinase dimer (227 microm). Enzymatic activity assay using the fluorescence resonance energy transfer method reveals that N-terminal deletion results in almost complete loss of enzymatic activity for SARS 3CL(pro). Molecular dynamics and docking simulations demonstrate the N-terminal deleted proteinase dimer adopts a state different from that of the full-length proteinase dimer, which increases the angle between the two protomers and reduces the binding pocket that is not beneficial to the substrate binding. This conclusion is verified by the surface plasmon resonance biosensor determination, indicating that the model substrate cannot bind to the N-terminal deleted proteinase. These results suggest the N terminus is not indispensable for the proteinase dimerization but may fix the dimer at the active state and is therefore vital to enzymatic activity.

Binding Sites↗

PubMed2004

MRI-based treatment planning for radiotherapy: dosimetric verification for prostate IMRT.

PURPOSE: Magnetic resonance (MR) and computed tomography (CT) image fusion with CT-based dose calculation is the gold standard for prostate treatment planning. MR and CT fusion with CT-based dose calculation has become a routine procedure for intensity-modulated radiation therapy (IMRT) treatment planning at Fox Chase Cancer Center. The use of MRI alone for treatment planning (or MRI simulation) will remove any errors associated with image fusion. Furthermore, it will reduce treatment cost by avoiding redundant CT scans and save patient, staff, and machine time. The purpose of this study is to investigate the dosimetric accuracy of MRI-based treatment planning for prostate IMRT. METHODS AND MATERIALS: A total of 30 IMRT plans for 15 patients were generated using both MRI and CT data. The MRI distortion was corrected using gradient distortion correction (GDC) software provided by the vendor (Philips Medical System, Cleveland, OH). The same internal contours were used for the paired plans. The external contours were drawn separately between CT-based and MR imaging-based plans to evaluate the effect of any residual distortions on dosimetric accuracy. The same energy, beam angles, dose constrains, and optimization parameters were used for dose calculations for each paired plans using a treatment optimization system. The resulting plans were compared in terms of isodose distributions and dose-volume histograms (DVHs). Hybrid phantom plans were generated for both the CT-based plans and the MR-based plans using the same leaf sequences and associated monitor units (MU). The physical phantom was then irradiated using the same leaf sequences to verify the dosimetry accuracy of the treatment plans. RESULTS: Our results show that dose distributions between CT-based and MRI-based plans were equally acceptable based on our clinical criteria. The absolute dose agreement for the planning target volume was within 2% between CT-based and MR-based plans and 3% between measured dose and dose predicted by the planning system in the physical phantom. CONCLUSIONS: Magnetic resonance imaging is a useful tool for radiotherapy simulation. Compared with CT-based treatment planning, MR imaging-based treatment planning meets the accuracy for dose calculation and provides consistent treatment plans for prostate IMRT. Because MR imaging-based digitally reconstructed radiographs do not provide adequate bony structure information, a technique is suggested for producing a wire-frame image that is intended to replace the traditional digitally reconstructed radiographs that are made from CT information.

Humans↗

PubMed2004

Multiplex mRNA assay using electrophoretic tags for high-throughput gene expression analysis.

We describe a novel multiplexing technology using a library of small fluorescent molecules, termed eTag molecules, to code and quantify mRNA targets. eTag molecules, which have the same fluorometric property, but distinct charge-to-mass ratios possess pre-defined electrophoretic characteristics and can be resolved using capillary electrophoresis. Coupled with primary Invader mRNA assay, eTag molecules were applied to simultaneously quantify up to 44 mRNA targets. This multiplexing approach was validated by examining a panel of inflammation responsive genes in human umbilical vein endothelial cells stimulated with inflammatory cytokine interleukin 1beta. The laser-induced fluorescence detection and electrokinetic sample injection process in capillary electrophoresis allows sensitive quantification of thousands of copies of mRNA molecules in a reaction. The assay is precise, as evaluated by measuring qualified Z' factor, a dimensionless and simple characteristic for applications in high-throughput screening using mRNA assays. Our data demonstrate the synergy between the multiplexing capability of eTag molecules by sensitive capillary electrophoresis detection and the isothermal linear amplification characteristics of the Invader assay. eTag multiplex mRNA assay presents a unique platform for sensitive, high sample throughput and multiplex gene expression analysis.

Cells, Cultured↗

PubMed2004

Stereotactic IMRT for prostate cancer: setup accuracy of a new stereotactic body localization system.

The purpose of this work is to prospectively assess the setup accuracy that can be achieved with a stereotactic body localizer (SBL) in immobilizing patients for stereotactic intensity-modulated radiotherapy (IMRT) for prostate cancer. By quantifying this important factor and target mobility in the SBL, we expect to provide a guideline for selecting planning target volume margins for stereotactic treatment planning. We analyzed data from 40 computed tomography (CT) studies (with slice thickness of 3 mm) involving 10 patients with prostate cancer. Each patient had four sets of CT scans during the course of radiotherapy. For the purpose of this study, all four sets of CT scans were obtained with the patients immobilized in a customized body pillow formed by vacuum suction. Unlike other immobilization devices, this system consists not only of a customized body pillow, but also of a fixation sheet used to suppress patient respiratory motion, a stereotactic body frame to provide stereotaxy, and a carbon fiber base board to which both the body cushion and the frame are affixed. We identified four bony landmarks and measured their coordinates in the stereotactic body frame on each set of CT scans. The displacements of the bony landmarks from their corresponding positions on the simulation scan (first CT scan) were analyzed in three dimensions in terms of overall, systematic, and random categories. The initial planned isocenter was also marked on the patients' skin with fiducials for each CT study. The distance from each bony landmark to the fiducial-based isocenter was measured and compared among the four sets of CT scans. The deviations in distances were also compared to those measured from the landmarks to the stereotactic frame center, in order to determine the effectiveness of the rigid body frame in positioning patients with prostate cancer. Target inter-fraction motion in this system was also studied for five patients by measuring the deviations in distances from the target geometric center to the bony landmarks. Our results showed that the overall setup accuracy had standard deviations (SDs) of 2.58 mm, 2.41 mm, and 3.51 mm in lateral (LAT), anterior-posterior(AP), and superior-inferior (SI) directions, respectively. The random component had SDs of 1.72 mm, 2.06 mm, and 2.79 mm, and the systematic component showed SDs of 0.92 mm, -0.27 mm, and 0.90 mm in these three directions. In terms of three-dimensional vector, the mean displacement over 116 measurements was 3.0 mm with an SD of 1.29 mm. Compared to the rigid reference, the skin-mark-based reference was less reliable for patient repositioning in terms of reproducing known bony landmark positions. The mean target mobility relative to the bony landmarks was 2.22 +/- 3.45 mm, 0.17 +/- 1.11 mm, and 0.11 +/- 2.69 mm in the AP, LAT, and SI directions, respectively. In conclusion, the body immobilization system has the ability to immobilize prostate cancer patients with satisfactory setup accuracy for fractionated extracranial stereotactic radiotherapy. A rigid frame system serves as a reliable alignment reference in terms of repositioning patients into the planning position, while skin-based reference showed larger deviations in repositioning patients.

Equipment Design↗

PubMed2004

Role of heart-type fatty acid binding protein in early detection of acute myocardial infarction in comparison with cTnI, CK-MB and myoglobin.

Heart fatty acid-binding protein (H-FABP) is supposed to be the most sensitive biomarker of early acute myocardial infarction (AMI). To evaluate the diagnostic value of H-FABP for AMI in the early stage, the plasma levels of H-FABP were measured by sandwich ELISA in 93 patients with suspected AMI at admission within 6 h after onset of chest pain and 69 normal healthy subjects. The plasma concentrations of cardiac troponin-I (cTnI), creatine kinase-MB (CK-MB) and myoglobin (Mb) were assayed at the same time by using corpuscle chemiluminescence for those patients. The patients were classified as AMI group (n=32) and non-AMI group (n=61) retrospectively. The diagnostic validity was evaluated in terms of sensitivity, specificity and receiver operating characteristic (ROC) curve analysis. The results showed the cutoff value of H-FABP for AMI was 16.8 ng/ml, and its diagnostic sensitivity for AMI was 64.29% within 3 h and 84.38% within 6 h after onset of chest pain, and the diagnostic specificity for non-AMI was 100% within 3 h and 91.8% within 6 h. H-FABP had higher sensitivity than that of cTnI and CK-MB at all time points (P<0.05), whereas there was no significant difference in specificity among the four markers. But the area under the ROC curve of H-FABP was significantly greater than that of cTnI, CK-MB and Mb within 3 h. These results revealed that H-FABP possessed high diagnostic sensitivity and specificity for AMI in early stage, especially within 3 h after onset of persistent angina pectoris. In conclusion, H-FABP can be used as a sensitive marker for AMI in the early stage.

Angina Pectoris↗

PubMed2004

Binding analyses between Human PPARgamma-LBD and ligands.

The binding characteristics of a series of PPARgamma ligands (GW9662, GI 262570, cis-parinaric acid, 15-deoxy-Delta(12,14)-prostaglandin J(2), LY171883, indomethacin, linoleic acid, palmitic acid and troglitazone) to human PPARgamma ligand binding domain have been investigated for the first time by using surface plasmon resonance biosensor technology, CD spectroscopy and molecular docking simulation. The surface plasmon resonance biosensor determined equilibrium dissociation constants (KD values) are in agreement with the results reported in the literature measured by other methods, indicating that the surface plasmon resonance biosensor can assume a direct assay method in screening new PPARgamma agonists or antagonists. Conformational changes of PPARgamma caused by the ligand binding were detected by CD determination. It is interesting that the thermal stability of the receptor, reflected by the increase of the transition temperature (T(m)), was enhanced by the binding of the ligands. The increment of the transition temperature (DeltaT(m)) of PPARgamma owing to ligand binding correlated well with the binding affinity. This finding implies that CD could possibly be a complementary technology with which to determine the binding affinities of ligands to PPARgamma. Molecular docking simulation provided reasonable and reliable binding models of the ligands to PPARgamma at the atomic level, which gave a good explanation of the structure-binding affinity relationship for the ligands interacting with PPARgamma. Moreover, the predicted binding free energies for the ligands correlated well with the binding constants measured by the surface plasmon resonance biosensor, indicating that the docking paradigm used in this study could possibly be employed in virtual screening to discover new PPARgamma ligands, although the docking program cannot accurately predict the absolute ligand-PPARgamma binding affinity.

Apoproteins↗

PubMed2003

Critical amino acid residues determine the binding affinity and the Ca2+ release efficacy of maurocalcine in skeletal muscle cells.

Maurocalcine (MCa) is a 33 amino acid residue peptide toxin isolated from the scorpion Scorpio maurus palmatus. MCa and mutated analogues were chemically synthesized, and their interaction with the skeletal muscle ryanodine receptor (RyR1) was studied on purified RyR1, sarcoplasmic reticulum (SR) vesicles, and cultured myotubes. MCa strongly potentiates [3H]ryanodine binding on SR vesicles (7-fold at pCa 5) with an apparent EC50 of 12 nm. MCa decreases the sensitivity of [3H]ryanodine binding to inhibitory high Ca2+ concentrations and increases it to the stimulatory low Ca2+ concentrations. In the presence of MCa, purified RyR1 channels show long-lasting openings characterized by a conductance equivalent to 60% of the full conductance. This effect correlates with a global increase in Ca2+ efflux as demonstrated by MCa effects on Ca2+ release from SR vesicles. In addition, we show for the first time that external application of MCa to cultured myotubes produces a cytosolic Ca2+ increase due to Ca2+ release from 4-chloro-m-cresol-sensitive intracellular stores. Using various MCa mutants, we identified a critical role of Arg24 for MCa binding onto RyR1. All of the other MCa mutants are still able to modify [3H]ryanodine binding although with a decreased EC50 and a lower stimulation efficacy. All of the active mutants produce both the appearance of a subconductance state and Ca2+ release from SR vesicles. Overall, these data identify some amino acid residues of MCa that support the effect of this toxin on ryanodine binding, RyR1 biophysical properties, and Ca2+ release from SR.

Amino Acid Sequence↗

PubMed2003

Maurocalcine and peptide A stabilize distinct subconductance states of ryanodine receptor type 1, revealing a proportional gating mechanism.

Maurocalcine (MCa) isolated from Scorpio maurus palmatus venom shares 82% sequence identity with imperatoxin A. Both scorpion toxins are putative mimics of the II-III loop peptide (termed peptide A (pA)) of alpha(1s)-dihydropyridine receptor and are thought to act at a common site on ryanodine receptor type 1 (RyR1) important for skeletal muscle EC coupling. The relationship between the actions of synthetic MCa (sMCa) and pA on RyR1 were examined. sMCa released Ca(2+) from SR vesicles (EC(50) = 17.5 nm) in a manner inhibited by micromolar ryanodine or ruthenium red. pA (0.5-40 microm) failed to induce SR Ca(2+) release. Rather, pA enhanced Ca(2+) loading into SR and fully inhibited Ca(2+)-, caffeine-, and sMCa-induced Ca(2+) release. The two peptides modified single channel gating behavior in distinct ways. With Cs(+)-carrying current, 10 nm to 1 microm sMCa induced long lived subconductances having 48% of the characteristic full open state and occasional transitions to 29% at either positive or negative holding potentials. In contrast, pA stabilized long lived channel closures with occasional burst transitions to 65% (s1) and 86% (s2) of the full conductance. The actions of pA and sMCa were observed in tandem. sMCa stabilized additional subconductance states proportional to pA-induced subconductances (i.e. 43% of pA-modified s1 and s2 substates), revealing a proportional gating mechanism. [(3)H]Ryanodine binding and surface plasmon resonance analyses indicated that the peptides did not interact by simple competition for a single class of mutually exclusive sites on RyR1 to produce proportional gating. The actions of sMCa were also observed with ryanodine-modified channels and channels deficient in immunophilin 12-kDa FK506-binding protein. These results provide evidence that sMCa and pA stabilize distinct RyR1 channel states through distinct mechanisms that allosterically stabilize gating states having proportional conductance.

Amino Acid Sequence↗

PubMed2003

One scorpion, two venoms: prevenom of Parabuthus transvaalicus acts as an alternative type of venom with distinct mechanism of action.

Scorpion venom is a complex mixture of salts, small molecules, peptides, and proteins. Scorpions employ this valuable tool in several sophisticated ways for subduing prey, deterring predators, and possibly during mating. Here, a subtle but clever strategy of venom utilization by scorpions is reported. Scorpions secrete a small quantity of transparent venom when initially stimulated that we propose to name prevenom. If secretion continues, a cloudy and dense venom that is white in color is subsequently released. The prevenom contains a combination of high K(+) salt and several peptides including some that block rectifying K(+) channels and elicit significant pain and toxicity because of a massive local depolarization. The presence of high extracellular K(+) in the prevenom can depolarize cells and also decrease the local electrochemical gradient making it more difficult to reestablish the resting potential. When this positive change to the K(+) equilibrium potential is combined with the blockage of rectifying K(+) channels, this further delays the recovery of the resting potential, causing a prolonged effect. We propose that the prevenom of scorpions is used as a highly efficacious predator deterrent and for immobilizing small prey while conserving metabolically expensive venom until a certain level of stimuli is reached, after which the venom is secreted.

Animals↗

PubMed2003

Diffusion-weighted MRI after cryosurgery of the canine prostate. Magnetic resonance imaging.

PURPOSE: To evaluate the acute lesion created by cryosurgery with diffusion-weighted magnetic resonance imaging (DWI). MATERIALS AND METHODS: The appearance of the acute cryolesion was evaluated in four canine prostates DWI after they were warmed to original body temperature. The prostates were excised, stained with triphenyl tetrazolium chloride (TTC), photographed, prepared for hematoxylin and eosin (H&E) staining, and examined under a light microscope. RESULTS: A marked decrease in apparent diffusion coefficient of 38% was evident in the center of the previously frozen tissue, but not in all of the previously frozen tissue. Histologic results confirm differences between the iceball core and the periphery of the iceball, which have markedly different imaging characteristics on DWI. CONCLUSION: The core of the previously frozen tissue has a reduced apparent diffusion coefficient (ADC) compared to the periphery of the previously frozen tissue and previously unfrozen tissue.

Animals↗

PubMed2002

A draft sequence of the rice genome (Oryza sativa L. ssp. japonica).

The genome of the japonica subspecies of rice, an important cereal and model monocot, was sequenced and assembled by whole-genome shotgun sequencing. The assembled sequence covers 93% of the 420-megabase genome. Gene predictions on the assembled sequence suggest that the genome contains 32,000 to 50,000 genes. Homologs of 98% of the known maize, wheat, and barley proteins are found in rice. Synteny and gene homology between rice and the other cereal genomes are extensive, whereas synteny with Arabidopsis is limited. Assignment of candidate rice orthologs to Arabidopsis genes is possible in many cases. The rice genome sequence provides a foundation for the improvement of cereals, our most important crops.

Arabidopsis↗

PubMed2002

Study of laser ablation in the in vivo rabbit brain with MR thermometry.

PURPOSE: To investigate the peak temperature and thermal dose (T(43)) as tissue damage indicators for thermal therapy. MATERIALS AND METHODS: The proton resonant frequency (PRF) shift thermal coefficient was calibrated on six in vivo rabbit brains during interstitial laser ablation. The peak temperature and T(43) were correlated with the lesion boundary observed on T2-weighted spin-echo (SE) MRI at 4 hours post-heating in seven thermal lesions using direct MR measurement and analysis based on a binary discriminate model. RESULTS: The peak temperature and T(43) were 48.3 +/- 1.7 degrees C and 191 +/- 219 minutes, respectively, from the direct MR measurement. The values derived by the binary discriminate analysis were 47.8 +/- 2.2 degrees C and 28 +/- 41 minutes, respectively. CONCLUSION: Our results suggest that tissue damage in rabbit brain 4 hours after thermal ablation can be predicted reliably from a threshold temperature of approximately 48 degrees C.

Animals↗

PubMed2002

[Extraction of ecdysterone from Stemmacantha uniflora].

OBJECTIVE: To optimize the conditions for the extraction of ecdysterone from Stemmacantha uniflora. METHODS: Conditions for the extraction were studied by three different methods as guided by the content of ecdysterone and solid material that present in the extract. RESULTS: Methanol was the best solvent, water the second, ethanol was greatly effected by concentration. Ultrasonic extraction had a good efficiency with short time. The time of cool extraction was too long, its efficiency was inferior to the other methods. CONCLUSION: The best methods for the extraction of ecdysterone from Stemmacantha uniflora was methanol reflux extraction.

Asteraceae↗