Biomedical subjects

Haiyan Liu

Publications and source records attributed to Haiyan Liu.

At least 19 recordsLinked to original sources

PubMed2026

Preclinical evaluation of AL-001, a gene therapy for wet age-related macular degeneration.

BACKGROUND: Frequent intravitreal administration of antivascular endothelial growth factor Vascular endothelial growth factor agents remains a major limitation in the management of wet age-related macular degeneration (wAMD). This study evaluated whether suprachoroidal delivery of an engineered recombinant adeno-associated viral (rAAV)-aflibercept vector could achieve sustained, targeted expression with improved efficacy and safety compared with intravitreal administration. METHODS: AL-001, an engineered rAAV vector expressing aflibercept, was developed and characterized. Its expression profile was first assessed in New Zealand white rabbits following suprachoroidal space (SCS) injection. Efficacy, pharmacokinetics, and safety were then evaluated in a nonhuman primate model of laser-induced choroidal neovascularization (CNV), comparing SCS and intravitreal (IVT) administration routes. RESULTS: AL-001 efficiently expressed aflibercept in relevant ocular cells in vitro. In rabbits, SCS administration produced sustained aflibercept levels in ocular tissues. In the nonhuman primate CNV model, a single SCS injection of AL-001 showed favorable efficacy to IVT injection and a notable mild inflammatory response. At week 4, grade IV lesion incidence was 0% (0/48) after SCS administration versus 14.3% (6/42) after IVT administration (absolute difference, -14.3 percentage points; 95% CI, 3.7%-27.8%; P = 0.0258). Throughout follow-up, mean leakage area and grade IV lesion incidence remained 0 with SCS, versus IVT peaks of approximately 0.3 mm2 and 33.0%, respectively, declining to 0.03 mm2 and 2.0% by day 100. Both the medium and high doses decreased pathological vascular leakage and subretinal hyperreflective material. Vector administration preceded laser-induced CNV modeling, demonstrating that sustained intraocular aflibercept expression in the retina and choroid provided durable antiangiogenic protection. Pharmacokinetic analysis confirmed distinct ocular exposure profiles between routes, with viral genomes confined predominantly to the injected eye and no significant systemic accumulation. AL-001 was well tolerated, without sustained intraocular pressure elevation or severe ocular inflammation, and only mild-to-moderate treatment-emergent adverse events. Low pre-existing anti-AAV2 immunity and time-dependent neutralizing antibody responses postdosing, informing a translational model for patient stratification and redosing feasibility. CONCLUSION: Suprachoroidal administration of AL-001 is well tolerated and provides durable, targeted aflibercept expression with pronounced antiangiogenic efficacy. These results support AL-001 as a promising, long-acting therapeutic candidate for wAMD.

AAV↗

PubMed2007

A comparative molecular dynamics study of thermophilic and mesophilic ribonuclease HI enzymes.

We studied a pair of homologous thermophilic and mesophilic ribonuclease HI enzymes by molecular dynamics simulations. Each protein was subjected to three 5 ns simulations in explicit water at both 310 K and 340 K. The thermophilic enzyme showed larger overall positional fluctuations at both temperatures, while only the mesophilic enzyme at the higher temperature showed significant instability. When the temperature is changed, the relative flexibility of different local segments on the two proteins changed differently. Principal component analysis showed that the simulations of the two proteins explored largely overlapping regions in the conformational space. However, at 340 K, the collective structure variations of the thermophilic protein are different from those of the mesophilic protein. Our results, although not in accordance with the view that hyperthermostability of proteins may originate from their conformational rigidity, are consistent with several recent experimental and simulation studies which showed that thermophilic proteins may be conformationally more flexible than their mesophilic counterparts. The decorrelation between conformational rigidity and hyperthermostability may be attributed to the temperature dependence and long range nature of electrostatic interactions that play more important roles in the structural stability of thermophilic proteins.

Computer Simulation↗

PubMed2006

A Bayesian-probability-based method for assigning protein backbone dihedral angles based on chemical shifts and local sequences.

Chemical shifts contain substantial information about protein local conformations. We present a method to assign individual protein backbone dihedral angles into specific regions on the Ramachandran map based on the amino acid sequences and the chemical shifts of backbone atoms of tripeptide segments. The method uses a scoring function derived from the Bayesian probability for the central residue of a query tripeptide segment to have a particular conformation. The Ramachandran map is partitioned into representative regions at two levels of resolution. The lower resolution partitioning is equivalent to the conventional definitions of different secondary structure regions on the map. At the higher resolution level, the alpha and beta regions are further divided into subregions. Predictions are attempted at both levels of resolution. We compared our method with TALOS using the original TALOS database, and obtained comparable results. Although TALOS may produce the best results with currently available databases which are much enlarged, the Bayesian-probability-based approach can provide a quantitative measure for the reliability of predictions.

Algorithms↗

PubMed2006

Impaired function and expression of P-glycoprotein in blood-brain barrier of streptozotocin-induced diabetic rats.

The aim was to investigate the effect of diabetes mellitus (DM) on P-glycoprotein (P-GP) function and expression in rat blood-brain barrier (BBB). P-GP function in BBB was assessed by measuring the brain-to-plasma concentration ratios (Kp values) of rhodamine 123 (Rho123) and vincristine (VCR), two well-known P-GP substrates, in control rats and 5-week streptozotocin (STZ)-induced diabetic rats. Evans blue (EB) dye was used as a BBB integrity indicator for examining the extravasation from the blood into the brain. P-GP expression in the brain cortex was evaluated with Western blot. The uptakes of Rho123 and VCR by cultured rat brain microvessel endothelial cells (rBMECs) incubated in diabetic and control rat serum for 72 h were also used to examine P-GP function, respectively. It was found that the Kp value of Rho123 (0.022+/-0.005 vs. 0.016+/-0.002 ml/g brain, p=0.033) and VCR (0.072+/-0.028 vs. 0.023+/-0.006 ml/g brain, p=0.006) in diabetic rats was significantly higher than that in control rats. The uptakes of Rho123 and VCR by cultured rBMECs incubated in the diabetic rat serum were higher than that in the control rat serum, respectively. No significant difference of the EB concentration in the brain cortex was found between the diabetic rats and control rats. Electron microscope examination of the brain cortex did not show a clear damage to the endothelial cells of microvessel in diabetic rats. In addition, the protein level of P-GP in the brains of the diabetic rats examined was significantly lower than that of control rats. These results suggested that the function and expression of P-GP might be impaired in the BBB of STZ-induced diabetic rats.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2006

QRAR models for cardiovascular system drugs using biopartitioning micellar chromatography.

The capability of biopartitioning micellar chromatography (BMC) to describe and estimate pharmacological parameters of cardiovascular system drugs has been studied. The retention of cardiovascular system drugs was studied using different pH of Brij-35 as micellar mobile phase in modified C(18) stationary phase. Quantitative retention-activity relationships (QRAR) in BMC were investigated for these compounds. An adequate correlation between the retention factors (log k) and the toxicity (LD(50)) of cardiovascular system drugs was obtained.

Cardiovascular Agents↗

PubMed2006

Binding model of human coactosin-like protein with filament actin revealed by mutagenesis.

Human coactosin-like protein (CLP) is a small (MW approximately 17 kDa) evolutionarily conserved actin-binding protein. It can bind to actin filaments but not globular actin and belongs to the fourth class of ADF-H-domain-containing proteins. Human CLP can also bind to 5LO, which plays an important role in cellular leukotriene synthesis. Although the structure of hCLP has been determined by both NMR and X-ray experiments, how hCLP binds to the actin filament is still a controversial question. To obtain insights into the structure of the complex, we studied the three-dimensional structure and backbone dynamics of hCLP using multidimensional NMR spectroscopy. Guided by the solution structure of the protein, a series of site-directed mutants were generated and their F-actin-binding activities were measured by high-speed cosedimentation assays. Furthermore, the structure model of the hCLP-F-actin complex was proposed using computational docking with the docking results filtered by the mutation data. Several previously untested residues (including T66, L89, R91, K102, D116 and E119) in hCLP were found important for the F-actin-binding activity. The extended region of beta4-beta5 of hCLP (residue 66-75) was found very flexible and very important for F-actin binding. The C-terminal residues of hCLP were not involved in F-actin binding, which was different from UNC-60B. Based on our hCLP-F-actin-binding model, different affinities of the four classes of ADF-H domain containing proteins for F-actin were explained.

Actins↗

PubMed2006

Determination of conformational free energies of peptides by multidimensional adaptive umbrella sampling.

We improve the multidimensional adaptive umbrella sampling method for the computation of conformational free energies of biomolecules. The conformational transition between the alpha-helical and beta-hairpin conformational states of an alanine decapeptide is used as an example. Convergence properties of the weighted-histogram-analysis-based adaptive umbrella sampling can be improved by using multiple replicas in each adaptive iteration and by using adaptive updating of the bounds of the umbrella potential. Using positional root-mean-square deviations from structures of the alpha-helical and beta-hairpin reference states as reaction coordinates, we obtained well-converged free energy surfaces of both the in-vacuum and in-solution decapeptide systems. From the free energy surfaces well-converged relative free energies between the two conformational states can be derived. Advantages and disadvantages of different methods for obtaining conformational free energies as well as implications of our results in studying conformational transitions of proteins and in improving force field are discussed.

Alanine↗

PubMed2006

Acidity adjustment of HZSM-5 zeolites by dealumination and realumination with steaming and citric acid treatments.

This article describes a novel method for acidity adjustment of HZSM-5 zeolites with steaming and citric acid treatments and demonstrates the realumination effect of citric acid on HZSM-5 zeolites dealuminated by steaming. A series of modified HZSM-5 zeolites were prepared by streaming and/or acid treatments and characterized by means of X-ray diffraction (XRD), X-ray fluorescence spectroscopy (XRF), (27)Al MAS NMR spectroscopy, hydroxyl infrared spectroscopy (OH-IR), pyridine-adsorbed infrared spectroscopy, and N(2) adsorption in the present investigation. The results showed that compared with single HCl or citric acid treatment, steaming treatment, and steaming/HCl treatments, citric acid treatment after steaming exclusively increased the amount of framework Al due to reinsertion of extraframework Al into the defective sites of the steamed HZSM-5 framework. This realumination effect of the citric acid treatment on the steamed HZSM-5 zeolite, which is reported here for the first time to the best of our knowledge, could nearly recover the pore structure of the steamed zeolite to that of the parent HZSM-5 zeolite and appropriately tailor the amount and strength of different acid sites, which sheds light on optimizing the physicochemical properties of HZSM-5 zeolites. It was also found that the steaming treatment prior to the citric acid treatment was the precondition of the realumination of HZSM-5 zeolites, suggesting that the lattice defect sites generated during steaming were necessary for citric acid to work.

Journal Article↗

PubMed2006

Identification of derlin-1 as a novel growth factor-responsive endothelial antigen by suppression subtractive hybridization.

Endothelial cells play an important regulatory role in embryonic development, reproductive functions, tumor growth and progression. In the present study, the suppression subtractive hybridization (SSH) method was employed to identify differentially expressed genes between non-stimulated endothelial cells and activated endothelial cells. Following mRNA isolation of non-stimulated and hepatocellular carcinoma homogenate-stimulated cells, cDNAs of both populations were prepared and subtracted by suppressive PCR. Sequencing of the enriched cDNAs identified a couple of genes differentially expressed, including derlin-1. Derlin-1 was significantly up-regulated by tumor homogenates, VEGF, and endothelial growth supplements in a dose-dependent manner. Knock-down of derlin-1 triggered endothelial cell apoptosis, inhibited endothelial cell proliferation, and blocked the formation of a network of tubular-like structures. Our data reveal that derlin-1 is a novel growth factor-responsive endothelial antigen that promotes endothelial cell survival and growth.

Antigens, Neoplasm↗

PubMed2006

On-line simultaneous removal of human serum albumin and enrichment of doxazosin using a weak cation-exchange monolithic column.

A weak cation-exchange monolithic column was prepared by modifying the GMA-EDMA (glycidyl methacrylate-co-ethylene glycol dimethacrylate) monoliths with ethylenediamine and monochloracetic acid. The properties of the column were investigated; the column exhibited the ability of low backpressure and fast analysis. Using this monolithic column, trace doxazosin in human serum albumin (HSA) solution and plasma samples has been on-line tested, the extraction efficiency and the maximum loading capacity of the monolithic column were obtained. The results showed that the monolithic column could realize deproteinization and trace drug enrichment simultaneously in the HSA solution and human plasma, which provided a simple, cheap, effective and friendly to environment method for assaying drugs in the blood.

Acetates↗

PubMed2006

Curcumin potently blocks Kv1.4 potassium channels.

Curcumin, a major constituent of the spice turmeric, is a nutriceutical compound reported to possess therapeutic properties against a variety of diseases ranging from cancer to cystic fibrosis. In whole-cell patch-clamp experiments on bovine adrenal zona fasciculata (AZF) cells, curcumin reversibly inhibited the Kv1.4K+ current with an IC50 of 4.4 microM and a Hill coefficient of 2.32. Inhibition by curcumin was significantly enhanced by repeated depolarization; however, this agent did not alter the voltage-dependence of steady-state inactivation. Kv1.4 is the first voltage-gated ion channel demonstrated to be inhibited by curcumin. Furthermore, these results identify curcumin as one of the most potent antagonists of these K+ channels identified thus far. It remains to be seen whether any of the therapeutic actions of curcumin might originate with its ability to inhibit Kv1.4 or other voltage-gated K+ channel.

Adrenal Glands↗

PubMed2006

Determination of metoprolol tartrate in tablets and human urine using flow-injection chemiluminescence method.

In this paper, a simple, rapid and sensitive flow-injection chemiluminescence method has been developed for the determination of metoprolol tartrate, which acts as a kind of sensitizer in the chemiluminescence emission from the redox of SO(3)(2-) with Ce(IV) in acidic medium. Under the optimized conditions, the proposed method allows the measurement of metoprolol tartrate over the range of 1.5 x 10(-8) to 7.3 x 10(-6)mol/L with a detection limit of 4.7 x 10(-9)mol/L (3sigma), and the relative standard deviation for 7.3 x 10(-7)mol/L metoprolol tartrate (n=11) is 2.20%. The utility of this method was demonstrated by determining metoprolol tartrate in tablets and human urine sample.

Cesium↗

PubMed2006

Molecular dynamics simulations of liquid methanol and methanol-water mixtures with polarizable models.

A polarizable model for simulation of liquid methanol, compatible with the COS/G2 water model, has been developed using the Charge-on-Spring (COS) technique. The model consists of three point charges, with one polarizable center on the oxygen atom. The Lennard-Jones parameters on the oxygen atom together with the molecular polarizability were varied to reproduce the experimental heat of vaporization and density of liquid methanol at ambient conditions. We examined the energies of various methanol dimers in the gas phase and compared them with values obtained from ab initio calculations. The model was then used to study the thermodynamic, dynamic, structural, and dielectric properties of liquid methanol as well as of a methanol-water mixture. A microscopic picture of the structure of pure liquid methanol and of the methanol-water mixture is provided. Good agreement was found between the results from our model simulations and available experimental and ab initio calculation data. In particular, the experimental dielectric permittivity of 32 could be reproduced, which had been shown to be difficult when using nonpolarizable models.

Dimerization↗

PubMed2006

Genetic algorithms for protein conformation sampling and optimization in a discrete backbone dihedral angle space.

We have investigated protein conformation sampling and optimization based on the genetic algorithm and discrete main chain dihedral state model. An efficient approach combining the genetic algorithm with local minimization and with a niche technique based on the sharing function is proposed. Using two different types of potential energy functions, a Go-type potential function and a knowledge-based pairwise potential energy function, and a test set containing small proteins of varying sizes and secondary structure compositions, we demonstrated the importance of local minimization and population diversity in protein conformation optimization with genetic algorithms. Some general properties of the sampled conformations such as their native-likeness and the influences of including side-chains are discussed.

Algorithms↗

PubMed2006

Abnormal activation of the synuclein-gamma gene in hepatocellular carcinomas by epigenetic alteration.

Liver cancer is the fifth most common neoplastic disease and the fourth leading cause of cancer-related death. Identification of the key molecular targets involved in hepatocarcinogenesis has significant therapeutic implications. In this study, by conducting immunohistochemistry, we show that the neuronal protein, synuclein-gamma (SNCG), is abnormally expressed in a high percentage of liver cancer (46/70, 65.7%). The expression of SNCG in liver cancer exhibits a clear stage-specific pattern of low expression in stage I (1/19, 5.3%) and high expression in stages III to IV (44/50, 88%). Importantly, all patients with metastatic diseases expressed SNCG in their primary tumors (15/15, 100%). Consistent with the IHC results, RT-PCR assays demonstrate that SNCG mRNA is highly expressed in the tumor tissue of advanced hepatocellular carcinomas. Analyses of the methylation status of the CpG island of the SNCG gene by methylation-specific PCR confirmed that all tumor samples contained the demethylated gene. To determine whether demethylation of SNCG is an early event of genetic abnormality in the process of hepatocarcinogenesis, we examined the methylation status of SNCG in 70 non-malignant cirrhotic liver samples and showed that 64.3% cirrhotic liver samples contained the partially or completely demethylated gene. We further show that SNCG expression in liver cancer is not restricted to HBV- and HCV-infected tumors, implying the involvement of other hepatocarcinogenic risk factors in SNCG gene reactivation. Utilizing human hepatoma-derived cell line HepG2 as an in vitro model, we demonstrate that hepatic carcinogens aflatoxin B1 and N-nitrosodimethylamine (DMN) are strong inducers of SNCG expression. Collectively, these new findings suggest that SNCG protein expression in primary tumors is a strong indicator of distant metastasis and demethylation of SNCG CpG island is an early sign of genetic abnormality in liver cirrhosis preceding hepatocarcinogenesis. Our studies also suggest that inducing demethylation of SNCG by hepatocarcinogens may represent one underlying mechanism for the aberrant expression of SNCG in HCC.

Adult↗

PubMed2005

Transition state ensemble for the folding of B domain of protein A: a comparison of distributed molecular dynamics simulations with experiments.

Folding pathways of the B domain of staphylococcal protein A have been sampled with a distributed computing approach. Starting from an extended structure, the method employs an index measuring topological similarity to the native structure to selectively sample trajectory branches leading to the native fold. Unperturbed and continuous folding trajectories are drawn on a physics-based atomic potential energy surface with an implicit solvent. The sampled folding trajectories demonstrate a similar sequence of events: the earlier stage involves a partial formation of helix 2 and to a less extent of helix 1 at their N terminals, followed by the hydrophobic collapse between residues F14, I17, and L18 on helix 1 and residues R28, F31, and I32 on helix 2, which results in the rigidification of the helix turn from R28 to I32. Helix 2 is then able to extend, allowing for the formation to turn 2. The above description explains one experimental result why a G30A mutant of the protein was observed to be the fastest folder among proteins of its size. And the ensemble of structures right before the final collapse is in good agreement with the transition state ensemble mapped by another recent experiment with Fersht Phi values. We emphasize that because the approach here does not provide quantifications of the free energy landscape, our model of the transition state ensemble emerges from comparisons of simulations and previous experimental results rather from the simulation results alone. On the other hand, as our approach does not rely on a low-dimensional free energy surface, it can complement methods based on the construction of free energy surfaces.

Amino Acid Sequence↗

PubMed2005

Structural basis for SUMO-E2 interaction revealed by a complex model using docking approach in combination with NMR data.

The interaction between small ubiquitin-related modifier SUMO and its conjugating-enzyme Ubc9 (E2) is an essential step in SUMO conjugation cascade. However, an experimental structure of such a transient complex is still unavailable. Here, a structural model of SUMO-3-Ubc9 complex was obtained with HADDOCK, combining NMR chemical shift mapping information. Docking calculations were performed using SUMO-3 and Ubc9 structures as input. The resulting complex reveals that the complementary surface electrostatic potentials contribute dominantly to the specific interaction. At the interface, similar numbers of oppositely-charged conserved residues are identified on the respective binding partners. Hydrogen bonds are formed in the vicinity of the interface to stabilize the complex. Comparison of the structure of SUMO-3-Ubc9 complex generated by HADDOCK and the experimental structures in free form indicates that SUMO-3 and Ubc9 maintain their respective fold as a whole after docking. However, the N-terminal helix alpha1 and its subsequent L1 loop of Ubc9 experience sizeable changes upon complex formation. They cooperatively move towards the hydrophilic side of the beta-sheet of SUMO-3. Our observations are consistent with the data from previous Ubc9 mutational analysis and conformational flexibility studies. Together, we have proposed that the SUMO-3-Ubc9 interaction is strongly electrostatically driven and the N terminus of Ubc9 shifts to SUMO-3 to facilitate the interaction. The NMR-based structural model, which provides considerable insights into the molecular basis of the specific SUMO-E2 recognition and interaction, implicates the general interaction mode between SUMO-3 and Ubc9 homologues from yeast to humans.

Binding Sites↗

PubMed2005

Loss of epigenetic control of synuclein-gamma gene as a molecular indicator of metastasis in a wide range of human cancers.

Metastasis is a major contributing factor to poor prognosis in cancer. Reliable and sensitive biomarkers that indicate the development of metastasis of primary tumors would be of great clinical use. In this study, we show that the neuronal protein synuclein-gamma (SNCG) is abnormally expressed in a high percentage (67.5%) of tumor tissues of diversified cancer types, including liver, esophagus, colon, gastric, lung, prostate, cervical, and breast cancer, but rarely expressed in tumor-matched nonneoplastic adjacent tissues (0.6%). Expressions of SNCG protein in different cancer types all display stage-specific patterns of very low expression in stage I and high expression in stages II to IV. Importantly, we observe a strong association between SNCG protein expression in primary tumors with distant metastasis in patients regardless of the cancer type (60.6%, P < 0.001). By performing genomic sequencing and methylation-specific PCR assays, we identify an inclusive demethylation of CpG sites within the CpG island of SNCG gene in every tumor sample (100%) across all cancer types, illustrating a universal loss of the epigenetic control of SNCG gene expression in tumors and further demonstrating that the demethylation of SNCG CpG island is primarily responsible for the aberrant expression of SNCG protein in cancerous tissues. These new findings strongly suggest that reactivation of SNCG gene expression by DNA demethylation is a common critical contributing factor to malignant progression of many solid tumors and its expression in primary carcinomas is an effective molecular indicator of distant metastasis. Our studies also suggest that the methylation status of SNCG gene can be used as a sensitive molecular tool in early detections of tumorigenesis.

Aged↗