Biomedical subjects

Jian Tan

Publications and source records attributed to Jian Tan.

At least 19 recordsLinked to original sources

PubMed2007

Reactive oxygen species mediates the apoptosis induced by transforming growth factor beta(2) in human lens epithelial cells.

Transforming growth factor beta(2) (TGF-beta(2)), a growth regulator of human lens epithelial cells (HLECs), also regulates the death of these cells. Dose-response analysis showed that the TGF-beta(2) concentration needed to induce HLECs death (100 pg/ml) was 10 times that needed to inhibit growth in these cells (10 pg/ml). TGF-beta(2)-induced apoptosis in HLECs was preceded by an induction of reactive oxygen species (ROS) and a decrease in glutathione in the intracellular content, indicating that this factor induces oxidative stress in HLECs. Studies performed to analyze the levels of c-fos mRNA, a gene whose expression is modulated by the redox state, demonstrated that only high, apoptotic concentrations of TGF-beta(2) (100 pg/ml) produced an increase in the mRNA levels of this gene, the level of induction being similar to that found when cells were incubated in the presence of hydrogen peroxide. Finally, the cell death induced by TGF-beta(2) in HLECs was partially blocked by radical scavengers, which decreased the percentage of apoptotic cells, whereas these agents did not modify the growth-inhibitory effect elicited by TGF-beta(2) in these cells. The results presented in this paper provide evidence for the involvement of an oxidative process in the apoptosis elicited by TGF-beta(2) in HLECs.

Apoptosis↗

PubMed2006

The Slc35d3 gene, encoding an orphan nucleotide sugar transporter, regulates platelet-dense granules.

Platelet dense granules are lysosome-related organelles which contain high concentrations of several biologically important low-molecular-weight molecules. These include calcium, serotonin, adenine nucleotides, pyrophosphate, and polyphosphate, which are necessary for normal blood hemostasis. The synthesis of dense granules and other lysosome-related organelles is defective in inherited diseases such as Hermansky-Pudlak syndrome (HPS) and Chediak-Higashi syndrome (CHS). HPS and CHS mutations in 8 human and at least 16 murine genes have been identified. Previous studies produced contradictory findings for the function of the murine ashen (Rab27a) gene in platelet-dense granules. We have used a positional cloning approach with one line of ashen mutants to establish that a new mutation in a second gene, Slc35d3, on mouse chromosome 10 is the basis of this discrepancy. The platelet-dense granule defect is rescued in BAC transgenic mice containing the normal Slc35d3 gene. Thus, Slc35d3, an orphan member of a nucleotide sugar transporter family, specifically regulates the contents of platelet-dense granules. Unlike HPS or CHS genes, it has no apparent effect on other lysosome-related organelles such as melanosomes or lysosomes. The ash-Roswell mouse mutant is an appropriate model for human congenital-isolated delta-storage pool deficiency.

Animals↗

PubMed2006

Interaction of Hermansky-Pudlak Syndrome genes in the regulation of lysosome-related organelles.

Hermansky-Pudlak Syndrome (HPS) is a genetically heterogeneous disease caused by abnormalities in the synthesis and/or trafficking of lysosome-related organelles (LROs) including melanosomes, lamellar bodies of lung type II cells and platelet dense granules. At least 15 genes cause HPS in mice, with a significant number specifying novel subunits of protein complexes termed BLOCs (Biogenesis of Lysosome-related Organelles Complexes). To ascertain whether BLOC complexes functionally interact in vivo, mutant mice doubly or triply deficient in protein subunits of the various BLOC complexes and/or the AP-3 adaptor complex were constructed and tested for viability and for abnormalities of melanosomes, lung lamellar bodies and lysosomes. All mutants, including those deficient in all three BLOC complexes, were viable though the breeding efficiencies of multiple mutants involving AP-3 were severely compromised. Interactions of BLOC protein complexes with each other and with AP-3 to affect most LROs were apparent. However, these interactions were tissue and organelle dependent. These studies document novel biological interactions of BLOC and AP-3 complexes in the biosynthesis of LROs and assess the role(s) of HPS protein complexes in general health and physiology in mammals. Double and triple mutant HPS mice provide unique and practical experimental advantages in the study of LROs.

Animals↗

PubMed2005

Slc7a11 gene controls production of pheomelanin pigment and proliferation of cultured cells.

In mammals, >100 genes regulate pigmentation by means of a wide variety of developmental, cellular, and enzymatic mechanisms. Nevertheless, genes that directly regulate pheomelanin production have not been described. Here, we demonstrate that the subtle gray (sut) mouse pigmentation mutant arose by means of a mutation in the Slc7a11 gene, encoding the plasma membrane cystine/glutamate exchanger xCT [Kanai, Y. & Endou, H. (2001) Curr. Drug Metab. 2, 339-354]. A resulting low rate of extracellular cystine transport into sut melanocytes reduces pheomelanin production. We show that Slc7a11 is a major genetic regulator of pheomelanin pigment in hair and melanocytes, with minimal or no effects on eumelanin. Furthermore, transport of cystine by xCT is critical for normal proliferation, glutathione production, and protection from oxidative stress in cultured cells. Thus, we have found that the Slc7a11 gene controls the production of pheomelanin pigment directly. Cells from sut mice provide a model for oxidative stress-related diseases and their therapies.

Amino Acid Transport System y+↗

PubMed2005

Computer construction and analysis of protein models of the mutant gammaD-crystallin gene.

BACKGROUND: Gammad-crystallin plays an important role in human cataract formation. Being highly stable, gammaD-crystallin proteins are composed of two domains. In this study we constructed and analyzed protein models of the mutant gammaD-crystallin gene, which caused a special fasciculiform congenital cataract affecting a large Chinese family. METHODS: gammaD-crystallin protein structure was predicted by Swiss-Model software using bovine gammaD-crystallin as a template and Prospect software using human betab2-crystallin as a template. The models were observed with a Swiss-Pdb viewer. RESULTS: The mutant gammaD-crystallin structure predicted by the Swiss-Model software showed that proline23 was an exposed surface residue and P23T change made a decreased hydrogen bond distance between threonine23 and asparagine49. The mutant gammaD-crystallin structure predicted by the Prospect software showed that the P23T change exerted a significant effect on the protein's tertiary structure and yielded hydrogen bonds with aspartic acid21, asparagine24, asparagine49 and serine74. CONCLUSION: The mutant gammaD-crystallin gene has a significant effect on the protein's tertiary structure, supporting that alteration of gamma-crystallin plays an important role in human cataract formation.

Animals↗

PubMed2005

The influence of microchannels on neurite growth and architecture.

Microchannels were produced using a photolithographic technique to pattern polyimide walls (11 microm in height and 20-60 microm in width) onto a planar glass substrate. PC12 cells were seeded onto patterned surfaces. After 3 days of culture in NGF supplemented medium cells were viable and extended neurites. Culture in microchannels influenced the direction of neurite growth (theta Orientation) and the complexity of PC12 cell architecture including neurite length (L(Neurite)), the number of neurites emerging per cell (N(Neurites)), and the angle at which neurites emerged from the cell soma (theta Soma). In microchannels neurites oriented parallel to channel walls and the complexity of neuronal architecture was reduced. Both of these effects were strongest for cells located in channels 20-30 microm wide. Within each channel the magnitude of the effect on orientation and architecture was inversely proportional to the distance of the soma from the channel wall. Microtubule and actin filament mobility within the cytoplasm may underly effect on neurite orientation and cell architecture. By manipulating channel width the overall direction of neurite growth and the complexity of neuronal architecture was controlled. Results from these studies will be applied towards the development of biomaterials for microfluidic platforms and drug discovery studies and in neural regeneration research-two applications that would be significantly improved given the ability to control neurite orientation and the complexity of neuronal architecture.

Animals↗

PubMed2005

Improved cell adhesion and proliferation on synthetic phosphonic acid-containing hydrogels.

Hydrogels with tissue-like mechanical properties are highly attractive scaffolds for tissue engineering. In this study, copolymers containing vinyl phosphonic acid (VPA) and acrylamide (AM) were tested for their swelling, protein uptake in serum supplemented medium, and cell adhesion and proliferation. The swelling of the gels in serum containing culture medium increased with increasing VPA content. The presence of VPA also increased protein uptake of gels in medium; gels polymerized with more than 50% of VPA absorbed as much as 100 microg/cm(2) of protein, twice the amount absorbed by gels made with only acrylamide. The adhesion and growth of the three types of cells, NIH 3T3 fibroblast, osteoblast-like MG-63 and Saos-2, were significantly improved on the gels made with 50% or more VPA; the number of adherent Mg-63 cells increased three-fold while the growth rate increased four-fold. Similar results were obtained for Saos-2 and 3T3 cells. The adhesion and growth of the three cell types on gels with sufficient phosphonate content were at least comparable to, or even better than, that on commercially available tissue culture plates. These results suggest great potential of anionic gels in bone tissue engineering.

Absorbable Implants↗

PubMed2004

Conformational searches elucidate effects of stereochemistry on structures of deoxyadenosine covalently bound to tumorigenic metabolites of benzo[C] phenanthrene.

Remarkably different conformations can result when DNA binds with stereoisomeric compounds containing differing absolute configurations of substituents about chiral carbon atoms. Furthermore, the biochemical functions of covalent adducts with DNA are strongly affected by the stereochemistry of the ligands. Such stereochemical effects are manifested by DNA covalent adducts derived from metabolites of the non-planar fjord region environmental chemical carcinogen benzo[c]phenanthrene. To analyze these phenomena, an extensive conformational investigation for R and S stereoisomeric adducts to deoxyadenosine, derived from trans addition of enantiomeric anti diol epoxide metabolites of benzo[c]phenanthrene, has been carried out. We have surveyed the potential energy surface of the two adducts by varying systematically at 5 degree intervals in combination, the three important torsion angles that govern conformational flexibility of the carcinogen bulk with respect to the linked nucleoside. We carried out a grid search by creating 373, 248 structures for each isomer, and evaluated their molecular mechanical energies. This has permitted us to map the potential energy surface of each adduct in these three variables, and to delineate their low energy regions. The maps have a symmetric relationship which stems from the near mirror-image stereochemistry in the R and S isomers. This produces near mirror-image low energy structures in the nucleoside adducts. The limited sets of stereoisomer-dependent conformational domains delineated are determined by steric effects. Moreover, these features have been experimentally demonstrated to play governing structural roles in such carcinogen-damaged DNA duplexes: opposite orientations in the stereoisomer pairs computed for the nucleosides are observed by high-resolution NMR in the similarly modified DNA double helices, and are likely to play important roles in their interactions with enzymes involved in DNA transactions, and hence their biological activities.

Carcinogens↗

PubMed2004

Surface-mediated gene transfer from nanocomposites of controlled texture.

Safe and efficient gene delivery would have great potential in gene therapy and tissue engineering, but synthetic biomaterial surfaces endowed with efficient gene-transferring functions do not yet exist. Inspired by naturally occurring biomineralization processes, we co-precipitated DNA with inorganic minerals onto cell-culture surfaces. The DNA/mineral nanocomposite surfaces obtained not only supported cell growth but also provided high concentrations of DNA in the immediate microenvironment of the cultured cells. Gene transfer from the engineered surfaces was as efficient as an optimized commercial lipid transfection reagent; in addition, the extent of gene transfer was adjustable by varying the mineral composition. DNA/mineral nanocomposite surfaces represent a promising system for enhancing gene transfer and controlling the extent of gene transfer for various biomedical applications, including tissue engineering or gene therapy of bone.

Animals↗

PubMed2004

Low power microwave radiation inhibits the proliferation of rabbit lens epithelial cells by upregulating P27Kip1 expression.

PURPOSE: The goal of this study was to examine the effects of low power microwave radiation (<10 mW/cm2) on the proliferation of cultured rabbit lens epithelial cells (RLEC). METHODS: Cultured RLEC were exposed to continuous microwave radiation at a frequency of 2,450 MHz and power densities of 0.10, 0.25, 0.50, 1.00, and 2.00 mW/cm2 for 8 h. Cell morphologic changes were observed under a phase-contrast microscope. Cell viability was measured using the MTT assay and cell cycle analysis was measured using flow cytometry. After exposure to 2.00 mW/cm2 microwave radiation for 4, 6, and 8 h, the expression of cell cycle-regulatory proteins, P21WAF1 and P27Kip1, was examined using western blot analysis. Finally, the levels of P21WAF1 and P27Kip1 mRNA were analyzed by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: After 8 h of radiation treatment, cells treated with 0.50, 1.00, and 2.00 mW/cm2 microwave radiation exhibited decreased cell viability, increased cell condensation and an inhibition of DNA synthesis. RLEC showed significant G0/G1 arrest. No obvious changes could be detected in the 0.10 and 0.25 mW/cm2 microwave treatment groups. Protein expression of P27Kip1 was markedly increased after microwave radiation. However, the mRNA levels were unchanged. On the other hand, there were no detectable differences in P21WAF1 protein expression and mRNA levels between microwave treatment and control groups. CONCLUSIONS: This study suggests that low power microwave radiation higher than 0.50 mW/cm2 can inhibit lens epithelial cell proliferation, and increase the expression of P27Kip1. These effects may account for the decline of lens epithelial proliferation after exposure to microwave radiation.

Animals↗

PubMed2004

The Hermansky-Pudlak syndrome 3 (cocoa) protein is a component of the biogenesis of lysosome-related organelles complex-2 (BLOC-2).

Hermansky-Pudlak syndrome (HPS) is a genetically heterogeneous inherited disease affecting vesicle trafficking among lysosome-related organelles. The Hps3, Hps5, and Hps6 genes are mutated in the cocoa, ruby-eye-2, and ruby-eye mouse pigment mutants, respectively, and their human orthologs are mutated in HPS3, HPS5, and HPS6 patients. These three genes encode novel proteins of unknown function. The phenotypes of Hps5/Hps5,Hps6/Hps6 and Hps3/Hps3,Hps6/Hps6 double mutant mice mimic, in coat and eye colors, in melanosome ultrastructure, and in levels of platelet dense granule serotonin, the corresponding phenotypes of single mutants. These facts suggest that the proteins encoded by these genes act within the same pathway or protein complex in vivo to regulate vesicle trafficking. Further, the Hps5 protein is destabilized within tissues of Hps3 and Hps6 mutants, as is the Hps6 protein within tissues of Hps3 and Hps5 mutants. Also, proteins encoded by these genes co-immunoprecipitate and occur in a complex of 350 kDa as determined by sucrose gradient and gel filtration analyses. Together, these results indicate that the Hps3, Hps5, and Hps6 proteins regulate vesicle trafficking to lysosome-related organelles at the physiological level as components of the BLOC-2 (biogenesis of lysosome-related organelles complex-2) protein complex and suggest that the pathogenesis and future therapies of HPS3, HPS5, and HPS6 patients are likely to be similar. Interaction of the Hps5 and Hps6 proteins within BLOC-2 is abolished by the three-amino acid deletion in the Hps6(ru) mutant allele, indicating that these three amino acids are important for normal BLOC-2 complex formation.

Alleles↗

PubMed2004

Biomaterials with hierarchically defined micro- and nanoscale structure.

Biomaterials and tissue engineering are becoming increasingly important in biomedical practice, particularly as the population ages. It is clear that cellular responses to materials depend on structural properties of the material at both the micrometer- and nanometer scale, but general methods for controlling material properties on both of these scales are lacking. Using a hierarchical approach that mimics natural material formation processes, we developed a method to produce materials with controlled physical structures at both the micrometer- and nanometer scale. Our method is based upon a pre-organized micropatterned template and conformal transformation of the architecture with nanostructured minerals, namely hydroxyapatite. The newly developed materials were biocompatible with bone cells, induced a range of desirable cellular responses, and may therefore have direct application in bone tissue engineering. In addition, the design principles employed in this study can be extrapolated to the other classes of biomedical materials, including polymers, metals, ceramics or hybrid combinations.

Biocompatible Materials↗

PubMed2004

Gels and more gels: probing toxicity.

Two-dimensional electrophoresis (2-DE) remains an important technology in the study of protein expression. In many applications, 2-DE has been supplanted by liquid chromatographic and mass spectrometric approaches that overcome some of its limitations and labor intensiveness. Nevertheless, 2-DE has exceptional relevance in toxicology and, despite the challenges, its implementation continues to support toxicologists in understanding the biological effects of chemical exposures in living systems. This review will address the use of 2-DE-based proteomics in toxicology and recent developments in this strategy, considering the unique nature of toxicity testing and the issues of dynamic range and reproducibility that have previously limited the overall utility of 2-DE in this field.

Animals↗

PubMed2004

[Low-intensity microwave blockes cell cycle and regulate cell cycle related gene expression in rabbit lens epithelial cells].

OBJECTIVE: To study the cell cycle arrest induced by different doses of microwave in cultured rabbit lens epithelial cells (RLECs) and its effects on the related gene expression of P21(WAF1), P27(Kip1) and c-myc. METHODS: Cultured rabbit lens epithelial cells were exposed to microwave radiation with frequency of 2450 MHz and power density of 0.10 mW/cm(2) (A group), 0.25 mW/cm(2) (B group), 0.50 mW/cm(2) (C group), 1.00 mW/cm(2) (D group) and 2.00 mW/cm(2) (E group) for 8 hours. The morphological change of RLECs was observed under microscope, effects of microwave radiation on RLECs cell cycle were measured by flow cytometry. Expression of P21(WAF1), P27(Kip1) and c-myc were evaluated by western blot after 2.00 mW/cm(2) microwave radiation for 4 hours, 6 hours and 8 hours respectively. All the results were compared with F group, which was as controls without microwave radiation. RESULTS: 8 hours after radiation, RLECs of C, D and E groups showed RLECs cell swelling, congregation and detachment, the percentage of cells in G(0)/G(1) phase obviously was increased (P < 0.01). No obvious change could be detected in 0.10 mW/cm(2) and 0.25 mW/cm(2) microwave groups. The gene expression of P27(Kip1) was increased and c-myc was decreased after 2.00 mW/cm(2) radiation, it did not affect the gene expression of P21(WAF1). CONCLUSION: Microwave exceeding 0.50 mW/cm(2) can block RLECs cell cycle at G(0)/G(1) phase, which may be related to regulation of P27(Kip1) and c-myc expression, but not P21(WAF1).

Animals↗

PubMed2003

Role of Crk-associated substrate in the regulation of vascular smooth muscle contraction.

A pool of actin monomers is induced to polymerize into actin filaments during contractile stimulation of smooth muscle. The inhibition of actin dynamics by actin polymerization inhibitors depresses active force generation in smooth muscle. In this study, we hypothesized that Crk-associated substrate plays a role in the regulation of contraction and actin dynamics in vascular smooth muscle. Antisense or sense oligodeoxynucleotides for Crk-associated substrate were introduced into carotid smooth muscle tissues by chemical loading. The treatment of smooth muscle strips with antisense oligodeoxynucleotides inhibited the expression of Crk-associated substrates; it did not influence the expression of actin, myosin heavy chain, and paxillin. Sense oligodeoxynucleotides did not affect the expression of these proteins in smooth muscle tissues. Force generation in response to stimulation with norepinephrine or KCl was significantly lower in antisense-treated muscle strips than in sense-treated strips or in muscle strips not treated with oligodeoxynucleotides. The downregulation of Crk-associated substrate did not attenuate increases in phosphorylation of the 20-kDa regulatory light chain of myosin in response to stimulation with norepinephrine. The increase in F-actin/G-actin ratio during contractile stimulation was significantly inhibited in antisense-treated smooth muscle strips. Contractile activation of smooth muscle increased the association of profilin with actin monomers; the depletion of Crk-associated substrate inhibited the increases in the profilin-actin complex in response to contractile stimulation. These results suggest that Crk-associated substrate is a necessary molecule of signaling cascades that regulate active force generation in smooth muscle. This molecule may regulate actin dynamics in smooth muscle in response to contractile stimulation.

Actins↗

PubMed2003

Downregulation of profilin with antisense oligodeoxynucleotides inhibits force development during stimulation of smooth muscle.

The actin-regulatory protein profilin has been shown to regulate the actin cytoskeleton and the motility of nonmuscle cells. To test the hypothesis that profilin plays a role in regulating smooth muscle contraction, profilin antisense or sense oligodeoxynucleotides were introduced into the canine carotid smooth muscle by a method of reversible permeabilization, and these strips were incubated for 2 days for protein downregulation. The treatment of smooth muscle strips with profilin antisense oligodeoxynucleotides inhibited the expression of profilin; it did not influence the expression of actin, myosin heavy chain, and metavinculin/vinculin. Profilin sense did not affect the expression of these proteins in smooth muscle tissues. Force generation in response to stimulation with norepinephrine or KCl was significantly lower in profilin antisense-treated muscle strips than in profilin sense-treated strips or in muscle strips not treated with oligodeoxynucleotides. The depletion of profilin did not attenuate increases in phosphorylation of the 20-kDa regulatory light chain of myosin (MLC20) in response to stimulation with norepinephrine or KCl. The increase in F-actin/G-actin ratio during contractile stimulation was significantly inhibited in profilin-deficient smooth muscle strips. These results suggest that profilin is a necessary molecule of signaling cascades that regulate carotid smooth muscle contraction, but that it does not modulate MLC20 phosphorylation during contractile stimulation. Profilin may play a role in the regulation of actin polymerization or organization in response to contractile stimulation of smooth muscle.

Actins↗

PubMed2003

Aberrant lung structure, composition, and function in a murine model of Hermansky-Pudlak syndrome.

Hermansky-Pudlak syndrome (HPS) is a genetically heterogeneous inherited disease causing hypopigmentation and prolonged bleeding times. An additional serious clinical problem of HPS is the development of lung pathology, which may lead to severe lung disease and premature death. No cure for the disease exists, and previously, no animal model for the HPS lung abnormalities has been reported. A mouse model of HPS, which is homozygously recessive for both the Hps1 (pale ear) and Hps2 (pearl) genes, exhibits striking abnormalities of lung type II cells. Type II cells and lamellar bodies of this mutant are greatly enlarged, and the lamellar bodies are engorged with surfactant. Mutant lungs accumulate excessive autofluorescent pigment. The air spaces of mutant lungs contain age-related elevations of inflammatory cells and foamy macrophages. In vivo measurement of lung hysteresivity demonstrated aberrant lung function in mutant mice. All these features are similar to the lung pathology described in HPS patients. Morphometry of mutant lungs indicates a significant emphysema. These mutant mice provide a model to further investigate the lung pathology and therapy of HPS. We hypothesize that abnormal type II cell lamellar body structure/function may predict future lung pathology in HPS.

Adaptor Protein Complex 3↗

PubMed2003

[Effect of low-intensity microwave radiation on proliferation of cultured epithelial cells of rabbit lens].

OBJECTIVE: To compare the effects of different doses of microwave on the proliferative activity and cell cycle of cultured epithelial cells of rabbit lens, and to investigate the limit tolerant of microwave exposure. METHODS: Cultured epithelial cells of rabbit lens were exposed to microwave radiation with frequency of 2,450 MHz and power density of 0.10, 0.25, 0.50, 1.00, 2.00 mW/cm(2) for 8 h in vitro. HE staining was used to observe the morphological changes of lens epithelial cells, the proliferative activity and cell cycle were measured by MTT assay and PI fluorescent staining. RESULTS: 8 h after radiation, 0.50, 1.00 and 2.00 mW/cm(2) microwave could decrease the proliferation of lens epithelial cells, make the cells disordered arrangement, shrinkage, detachment, and inhibit the synthesis of cell DNA. The percentage of G(0)/G(1) phase cells were 71.95% +/- 2.12%, 75.68% +/- 3.35% and 82.40% +/- 8.68% respectively, which were higher than that in control group (61.68% +/- 5.76%, P < 0.05 or P < 0.01). The percentage of S phase cells were 19.32% +/- 3.07%, 16.08% +/- 4.91% and 12.98% +/- 8.08% respectively, which were lower than that in control group (28.05% +/- 5.12%, P < 0.05 or P < 0.01). No obvious changes could be detected in 0.10, 0.25 mW/cm(2) microwave groups (P > 0.05). CONCLUSION: Microwave exceeding 0.50 mW/cm(2) may make injury to lens epithelial cells after 8 hour radiation, which may be related to the effect of microwave radiation on cell cycle.

Animals↗