Biomedical subjects

Jian Shen

Publications and source records attributed to Jian Shen.

At least 55 records · Page 3Linked to original sources

PubMed2003

Argyrophilic nuclear organizer regions in nasopharyngeal carcinoma and paraneoplastic epithelia.

BACKGROUND: Nucleolar organizer regions (NORs), which are loops of DNA containing ribosomal RNA genes, have been shown to correlate with cell proliferation and malignant transformation. The diagnostic value of silver staining NORs (AgNORs) in preneoplastic epithelia and nasopharyngeal carcinoma (NPC) by the quantitative assessment of AgNORs proteins is evaluated. METHODS: Silver-staining of NORs was applied to 70 paraffin sections of NPC biopsy specimens, including 29 samples of the adjacent normal glandular epithelia (GE), 19 hyperplastic/dysplastic columnar epithelia (CE) in the adjacent mucosa, 10 hyperplastic/dysplastic squamous epithelia (SE) in the adjacent mucosa, 54 differentiated nonkeratinizing carcinoma (DC), and 16 undifferentiated carcinoma (UC). The morphometric features of AgNORs in preneoplastic epithelia and cancer cells were analyzed by image cytometric analysis and then compared. RESULTS: The AgNOR count, mean AgNOR area, and AgNOR area-count ratio increased significantly from GE to CE to DC or UC. The mean nuclear area, AgNOR area, and AgNOR area-count ratio increased significantly from GE to SE to UC, whereas overlapping AgNOR values were observed between SE and DC. UC had significantly higher values than DC in mean nuclear area, AgNOR area, and AgNOR area-count ratio. CONCLUSIONS: The morphometric analysis of AgNORs reflected cell proliferative activity and histologic differentiation of tumor rather than malignant transformation in different nasopharyngeal epithelia and NPC. AgNOR area and AgNOR area-count ratio are the most valuable features for differential diagnosis of normal, preneoplastic, and cancer cells.

Adult↗

PubMed2003

Platelet adhesive resistance of segmented polyurethane film surface-grafted with vinyl benzyl sulfo monomer of ammonium zwitterions.

Platelet from human plasma adhered on the segmented poly(ether urethane) (SPEU) film grafted with N,N-dimethyl-N-(p-vinylbenyl)-N-(3-sulfopropyl) ammonium (DMVSA) was studied. SPEU films were hydroxylated by potassium peroxosulfate (KPS) and then grafted with DMVSA using ceric ammonium nitrate (CAN) as initiator. The mixing time of hydroxylated SPEU/CAN and the monomer concentration effect on graft polymerization yield were determined by ATR-FTIR. Surface analysis of the grafted films by ATR-FTIR and ESCA confirmed that DMVSA was successfully grafted onto the SPEU film surface. The grafted film possessed a relatively hydrophilic surface, as revealed by water contact angle measurement. The improved blood compatibility of the grafted films was preliminarily evaluated by a platelet-rich plasma adhesion study and scanning electron microscopy, using original SPEU and hydroxylated SPEU films as the controls. The results showed that platelet attachment was decreased greatly on the segmented polyurethane films grafted with DMVSA. This kind of new biomaterials grafted with sulfo ammonium zwitterionic monomers might have potential for biomedical applications.

Blood Platelets↗

PubMed2003

The Saccharomyces cerevisiae Arr4p is involved in metal and heat tolerance.

Homologues of the bacterial ArsA ATPase are found in nearly every organism. While the enzyme is involved in arsenic detoxification in bacteria, the roles of eukaryotic homologues have not been identified. This article reports the function of the Saccharomyces cerevisiae homologue encoded by ARR4 gene (YDL100c ORF). Disruption of ARR4 was not lethal, but the disrupted strain displayed increased sensitivity to As3+, As5+, Co2+, Cr3+, Cu2+ or VO4(3-) salts and temperature. A plasmid-encoded copy of a wild-type ARR4 gene could complement the heat- or metal-related stress responses. Mutation of a codon within the consensus sequence for the nucleotide-binding site resulted in loss of complementation of the disrupted strain and produced a dominant negative phenotype in a wild type strain. Wild type and mutant Arr4p were purified from Escherichia coli. The wild type protein exhibited a low level of ATPase activity, and the mutant was inactive. The purified ATPase eluted as a dimer of 80-kDa species. A fusion of ARR4 and the GFP (green fluorescent protein) gene was constructed. The gene fusion was able to complement stress-related phenotype of the ARR4 disruption. Under non-stress conditions, GFP fluorescence was found diffusely in the cytosol. Under stress conditions GFP was localized in a few punctate bodies resembling late endosomes. It is proposed that under heat or metal stress, the soluble ATPase becomes membrane-associated, perhaps through interaction with a partner protein, and that this complex is involved in stress tolerance.

Adaptation, Physiological↗

PubMed2003

Covalent immobilization of O-butyrylchitosan with a photosensitive hetero-bifunctional crosslinking reagent on biopolymer substrate surface and bloodcompatibility characterization.

O-Butyrylchitosan (OCS) was covalently immobilized onto a substrate (suture and PTFE) surface using the photosensitive hetero-bifunctional crosslinking reagent 4-azidobenzoic acid, which was previously bonded to OCS by the reaction between an acid group of the crosslinking reagent and a free amino group of OCS. The immobilization was accomplished by irradiating the modified OCS coated on the substrate surface with ultraviolet light to photolyze azide groups, thus the crosslinking OCS was immobilized on the substrate surface. The result indicated that OCS molecules immobilized on the substrate, and significantly reduced the fibrinogen adsorption and the deposition and spreading of platelets, demonstrating superior bloodcompatibility. Therefore, OCS could be developed into bloodcompatible biomaterial that used to modify the biomedical devices, such as PTFE and Dacron vascular prostheses surface.

Animals↗

PubMed2003

Reactive oxygen species and antioxidants in apoptosis of esophageal cancer cells induced by As2O3.

To explore the relationship between the reactive oxygen species (ROS) and apoptosis in esophageal carcinoma cells (SHEE85) induced by arsenic trioxide (As2O3), we focused on changes of apoptosis, ROS, and antioxidants. Apoptosis of SHEE85 was confirmed by means of DNA fragmentation stained by Hoechst 33342, Sub-G1 cells scored by flow cytometry and ultrastructure of cells by electron microscopy. To evaluate the level of ROS, the chemiluminescent method was used for measuring the production of superoxide anion (O(-)*2). Lipid peroxide (malondialdehyde, MDA), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) were measured respectively by the photometry method. In the cells treated with As2O3 at a concentration of 5.0 micromol/l for 2-24 h, the content of cellular O(-)*2 and MDA was increased, but SOD and GSH-Px were significantly lower in the process of apoptosis in SHEE85. As2O3 at concentration of 0.5 micromol/l did not cause cell apoptosis but promoted cell proliferation. These results suggest that As2O3 at a high dosage (5 micromol/l) causes cell apoptosis and at a low dosage (0.5 micromol/l) causes cell proliferation. The essential mechanisms of cell apoptosis induced by As2O3 may be related to the increase of ROS and decrease of anti-oxidation. ROS and antioxidants participate in the apoptotic pathway of esophageal carcinoma cells.

Antineoplastic Agents↗

PubMed2003

Cytogenetic and molecular genetic changes in malignant transformation of immortalized esophageal epithelial cells.

The purpose of this study was to evaluate the extent to which the expression of p53, c-myc, bcl-2, ras genes and chromosomes, along with activity of hTERT, impacts on the malignant transformation of immortalized esophageal epithelial cells. The SHEE cell line was established from an embryonic esophageal epithelial cell induced by transduction of E6E7 genes of human papillomavirus type 18 (HPV18E6E7). In cells of the 85th passage (SHEE85), the malignant transformation of SHEE was confirmed by morphology, cell proliferative index and tumor formation in SCID mice. C-myc, p53, bcl-2 and ras genes were assayed by the multi-PCR method with house-keeping gene GAPDH as control. The modal number of chromosomes was analyzed and its expression of subunit of telomerase, hTERT, was assessed by RT-PCR. Expression of HPV18E6E7 was assayed by Western blotting. The results showed that cells of SHEE85 were atypical and exhibited proliferative status with a proliferation index of 45.70%. Tumors formed in SCID mice with invasion of adjacent tissue. The karyotype belonged to hypotriploid and displayed expression of hTERT. C-myc, k-ras, bcl-2 and p53 (expression of phosphoprotein) were positive in SHEE85. Expression of HPV18E6E7 was positive. Taken together, SHEE85 cells were in fully malignant transformation and their molecular mechanism involved the expression of cellular genes, such as p53, bcl-2, c-myc and ras, and aberrance of chromosomes. It is probable that all of these changes were related with HPV18E6E7.

Animals↗

PubMed2003

Ezrin, actin and cytoskeleton in apoptosis of esophageal epithelial cells induced by arsenic trioxide.

Ezrin is a key protein in membrane-cytoskeleton interaction. Expression of ezrin in actin-rich cell surfaces may play a role in the modulation of cell shape and adhesion. The aim of this study was to detect ezrin, actin and cytoskeleton and to explore their relationship in the apoptosis of esophageal epithelial cells (SHEE) induced by arsenic trioxide (As2O3). The SHEE is an immortalized human fetal esophageal epithelial cell line, and the cells were treated by administering 5, 10 and 20 micromol of As2O3. the proliferation and apoptosis of SHEE cells were examined by flow cytometry with propidium iodide staining. Ezrin expression was detected by immunocytochemistry and Western blotting. Actin filament was stained by FITC-labeled phalloidin and detected quantitatively by fluorescent microscopy. Cell morphology and microfilaments were examined by electron microscopy. The results revealed that As2O3 induced an inhibition of proliferation and the promotion of apoptosis in SHEE cells. The ezrin, actin and cytoskeleton were decreased after As2O3 treatment and the cellular morphology of apoptosis developed. Our results suggested that the morphological changes of arsenic-induced apoptosis of human esophageal epithelial cells were initiated by ezrin and actin-cytoskeletal aberrance.

Actins↗

PubMed2003

[Development of the vascular prosthesis research].

The search for a nonthrombogenic material with the potential for use in small diameter vascular graft applications continues to be a field of extensive investigation. This article describes the choice of biomaterials used as vascular prosthesis, the innovation of construction of tissue-engineered blood vessels, the indispensability, methods and the effect produced by surface modification of vascular prosthesis. The article also points out that research achievements of vascular prosthesis must be made with the exploitation of new nonthrombogenic biomaterial and the development of tissue engineering.

Biocompatible Materials↗

PubMed2002

Self-assembly of nanometer-scale magnetic dots with narrow size distributions on an insulating substrate.

The self-assembly of iron dots on the insulating surface of NaCl(001) is investigated experimentally and theoretically. Under proper growth conditions, nanometer-scale magnetic iron dots with remarkably narrow size distributions can be achieved in the absence of a wetting layer. Furthermore, both the vertical and lateral sizes of the dots can be tuned with the iron dosage without introducing apparent size broadening, even though the clustering is clearly in the strong coarsening regime. These observations are interpreted using a phenomenological mean-field theory, in which a coverage-dependent optimal dot size is selected by strain-mediated dot-dot interactions.

Journal Article↗

PubMed2002

Arsenite transport by mammalian aquaglyceroporins AQP7 and AQP9.

Much is known about the transport of arsenite and antimonite into microbes, but the identities of mammalian transport proteins are unknown. The Saccharomyces cerevisiae FPS1 gene encodes a membrane protein homologous to the bacterial aquaglyceroporin GlpF and to mammalian aquaglyceroporins AQP7 and AQP9. Fps1p mediates glycerol uptake and glycerol efflux in response to hypoosmotic shock. Fps1p has been shown to facilitate uptake of the metalloids arsenite and antimonite, and the Escherichia coli homolog, GlpF, facilitates the uptake and sensitivity to metalloid salts. In this study, the ability of mammalian aquaglyceroporins AQP7 and AQP9 to substitute for the yeast Fps1p was examined. The fps1Delta strain of S. cerevisiae exhibits increased tolerance to arsenite and antimonite compared to a wild-type strain. Introduction of a plasmid containing AQP9 reverses the metalloid tolerance of the deletion strain. AQP7 was not expressed in yeast. The fps1Delta cells exhibit reduced transport of (73)As(III) or (125)Sb(III), but uptake is enhanced by expression of AQP9. Xenopus laevis oocytes microinjected with either AQP7 or AQP9 cRNA exhibited increased transport of (73)As(III). These results suggest that AQP9 and AQP7 may be a major routes of arsenite uptake into mammalian cells, an observation potentially of large importance for understanding the action of arsenite as a human toxin and carcinogen, as well as its efficacy as a chemotherapeutic agent for acute promyelocytic leukemia.

Animals↗

PubMed2002

Unisite and multisite catalysis in the ArsA ATPase.

The ars operon of plasmid R773 encodes an As(III)/Sb(III) extrusion pump. The catalytic subunit, the ArsA ATPase, has two homologous halves, A1 and A2, each with a consensus nucleotide-binding sequence. ATP hydrolysis is slow in the absence of metalloid and is accelerated by metalloid binding. ArsA M446W has a single tryptophan adjacent to the A2 nucleotide-binding site. Tryptophan fluorescence increased upon addition of ATP, ADP, or a nonhydrolyzable ATP analogue. Mg(2+) and Sb(III) produced rapid quenching of fluorescence with ADP, no quenching with a nonhydrolyzable analogue, and slow quenching with ATP. The results suggest that slow quenching with ATP reflects hydrolysis of ATP to ADP in the A2 nucleotide-binding site. In an A2 nucleotide-binding site mutant, nucleotides had no effect. In contrast, in an A1 nucleotide-binding mutant, nucleotides still increased fluorescence, but there was no quenching with Mg(2+) and Sb(III). This suggests that the A2 site hydrolyzes ATP only when Sb(III) or As(III) is present and when the A1 nucleotide-binding domain is functional. These results support previous hypotheses in which only the A1 nucleotide-binding domain hydrolyzes ATP in the absence of activator (unisite catalysis), and both the A1 and A2 sites hydrolyze ATP when activated (multisite catalysis).

Adenosine Triphosphate↗

PubMed2002

Tabtoxin-resistant protein: overexpression, purification, and characterization.

One of the self-protection mechanisms in Pseudomonas syringae pv. tabaci, a pathogen of tobacco wildfire, is thought to be due to its tabtoxin-resistance gene (ttr). In this study, the ttr gene was inserted into an expression vector, pQE30, and successfully expressed in Escherichia coli M15 at high levels. The purified recombinant tabtoxin-resistant protein (TTR) had an apparent molecular mass of about 21 kDa on SDS-PAGE as well as by mass spectroscopy and had a pI of 6.6 on isoelectric focusing-PAGE. Spectral analysis showed that TTR possesses a maximum fluorescence wavelength (lambda(max)) of 325 nm upon excitation at 282 nm and a positive band with a maximum at 195 nm and a broad negative band with a minimum at 215 nm in the far-UV CD spectrum. The spectrophotometric assay demonstrated the strong detoxification activity of TTR. These results are the first report of the characterization of the purified tabtoxin-resistant protein. Its capacity to detoxify tabtoxinine-beta-lactam shows that it must be one of the self-protection mechanisms in pv. tabaci.

Acetyltransferases↗

PubMed2002

Covalent immobilization of chitosan/heparin complex with a photosensitive hetero-bifunctional crosslinking reagent on PLA surface.

Chitosan (CS) was covalently immobilized onto polylactic acid (PLA) film surface using the photosensitive hetero-bifunctional crosslinking reagent, 4-azidobenzoic acid, which was previously bonded to chitosan by reaction between an acid group of the crosslinking reagent and a free amino group of chitosan. The immobilization was accomplished by irradiating with ultraviolet light, the modified chitosan being coated on the film surface to photolyze azide groups, thus crosslinking chitosan and PLA together. The hydroxyl and amino groups of chitosan may provide the opportunity for them being further derived by chemically immobilizing a variety of functional groups. These modifications could be carried out to tailor PLA biomaterial to meet the specific needs of different biomedical applications. For example, chitosan molecules immobilized on the PLA could be modified by heparin (Hp) solution to form a polyelectrolyte complex on the PLA surface. Platelet adhesion assay showed that PLA surface modified by chitosan/heparin complex could inhibit platelet adhesion and activation. Cell culture assay indicated that PLA surface with CS/Hp complex showed enhanced cell adhesion.

Animals↗

PubMed2002

Cell adhesion behavior of chitosan surface modified by bonding 2-methacryloyloxyethyl phosphorylcholine.

2-Methacryloyloxyethyl phosphorylcholine (MPC)-bonded chitosan was prepared by Michael addition of MPC to the amino groups of chitosan. The modified surfaces were characterized by static contact angle and electron spectroscopy for chemical analysis (ESCA). The water contact angle of chitosan decreased with the MPC bonding and the rate of decrease depended on the amount of MPC bonding. ESCA analysis results proved that MPC had been bonded on the chitosan surface and the chitosan modified directly by MPC had a much higher concentration of MPC on the surface compared with that of MPC on chitosan modified indirectly by MPC. Cell adhesion tests indicated that a low concentration of MPC bonded chitosan was more favorable to cell adhesion while a high concentration of MPC bonded chitosan inhibited cell attachment.

Cell Adhesion↗

PubMed2002

Morphological and functional changes of mitochondria in apoptotic esophageal carcinoma cells induced by arsenic trioxide.

AIM: To demonstrate that mitochondrial morphological and functional changes are an important intermediate link in the course of apoptosis in esophageal carcinoma cells induced by As2O3. METHODS: The esophageal carcinoma cell line SHEEC1, established in our laboratory, was cultured in 199 growth medium, supplemented with 100mL x L(-1) calf serum and 3 mol x L(-1)As2O3 (the same below). After 2, 4, 6, 12, 24 h of drug adding, the SHEEC1 cells were collected for light-and electron-microscopic examination. The mitochondria were labeled by Rhodamine fluorescence probe and the fluorescence intensity of the mitochondria was measured by flow cytometer and cytofluorimetric analysis. Further,the mitochondrial transmembrane potential (MTP, psim) change was also calculated. RESULTS: The mitochondrial morphological change after adding As2O3 could be divided into three stages. In the early-stage (2-6h) after adding As2O3, an adaptive proliferation of mitochondria appeared; in the mid-stage (6-12 h) a degenerative change was observed; and in the late-stage (12-24 h) the mitochondria swelled with outer membrane broken down and then cells death with apoptotic changes of nucleus. The functional change of the mitochondria indicated by fluorescent intensity, which reflected the MTP status of mitochondria, was in accordance with morphological change of the mitochondria. The fluorescent intensity increased at early-stage, declined in mid-stage and decreased to the lowest in the late-stage. 24 h after As2O3 adding, the cell nucleus showed typical apoptotic changes. CONCLUSION: Under the inducement of As2O3, the early apoptotic changes of SHEEC1 cells were the apparent morphological and functional changes of mitochondria, afterwards the nucleus changes followed. It is considered that changes of mitochondria are an important intermediate link in the course of apoptosis of esophageal carcinoma cells induced by As2O3.

Antineoplastic Agents↗

PubMed2002

Nitric oxide and calcium ions in apoptotic esophageal carcinoma cells induced by arsenite.

AIM: To Quantitatively analyze the nitric oxide (NO) and Ca2+ in apoptosis of esophageal carcinoma cells induced by arsenic trioxide (As2O3). METHODS: The cell line SHEEC1, a malignant esophageal epithelial cell induced by HPV in synergy with TPA in our laboratory, was cultured in a serum-free medium and treated with As2O3. Before and after administration of As2O3, NO production in cultured medium was detected quantitatively using the Griess Colorimetric method. Intracellular Ca2+ was labeled by using the fluorescent dye Fluo3-AM and detected under confocal laser scanning microscope (CLSM), which was able to acquire data in real-time enabling Ca2+ dynamics of individual cells in vitro. The apoptotic cells were examined under electron microscopy. RESULTS: Intracellular concentration of Ca2+ increased from 1.00 units to 1.09-1.38 units of fluorescent intensity at As2O3 treatment and NO products subsequently released from As2O3-treated cells increased from 0.98-1.00 x10(-2)micromol x L(-1) up to 1.48-1.52 x10(-2)micromol x L(-1) and maintained in a high level continuously. Finally apoptosis of cells occurred,chromatin being agglutinated, cells shrunk,nuclei became round and mitochondria swelled. CONCLUSION: Ca2+ and NO increased with cell damage and apoptosis in cells treated by As2O3. The Ca2+ is an initial messenger to the apoptotic pathway. To investigate Ca2+ and NO will be a new direction for studying the apoptotic signaling messenger of the esophageal carcinoma cells induced by As2O3.

Antineoplastic Agents↗

PubMed2002

Telomere and telomerase in the initial stage of immortalization of esophageal epithelial cell.

AIM: To search for the biomarker of cellular immortalization, the telomere length, telomerase activity and its subunits in cultured epithelial cells of human fetal esophagus in the process of immortalization. METHODS: The transgenic cell line of human fetal esophageal epithelium (SHEE) was established with E(6)E(7) genes of human papillomavirus (HPV) type 18 in our laboratory. Morphological phenotype of cultured SHEE cells from the 6th to 30th passages, was examined by phase contrast microscopy, the telomere length was assayed by Southern blot method, and the activity of telomerase was analyzed by telomeric repeat amplification protocol (TRAP). Expressions of subunits of telomerase, hTR and hTERT, were assessed by RT-PCR. DNA content in cell cycle was detected by flow cytometry. The cell apoptosis was examined by electron microscopy (EM) and TUNEL label. RESULTS: SHEE cells from the 6th to 10th passages showed cellular proliferation with a good differentiation. From the 12th to the 16th passages, many senescent and apoptotic cells appeared, and the telomere length sharply shortened from 23kb to 17kb without expression of hTERT and telomerase activity. At the 20th passage, SHEE cells overcame the senescence and apoptosis and restored their proliferative activity with expression of telomerase and hTERT at low levels, but the telomere length shortened continuously to the lowest of 3kb. After the 30th passage cells proliferation was restored by increment of cells at S and G2M phase in the cell cycle and telomerase activity expressed at high levels and with maintenance of telomere length. CONCLUSION: At the early stage of SHEE cells, telomeres are shortened without expression of telomerase and hTERT causing cellular senescence and cell death. From the 20th to the 30th passages, the activation of telomerase and maintenance of telomere length show a progressive process for immortalization of esophageal epithelial cells. The expression of telomerase may constitute a biomarker for detection of immortalization of cells.

Apoptosis↗

PubMed2002

Progressive transformation of immortalized esophageal epithelial cells.

AIM: To investigate the progressive transformation of immortal cells of human fetal esophageal epithelium induced by human papillomavirus, and to examine biological criteria of sequential passage of cells, including cellular phenotype, proliferative rate, telomerase, chromosome and tumorigenicity. METHODS: The SHEE cell series consisted of immortalized embryonic esophageal epithelium which was in malignant transformation when cultivated over sixty passages without co-carcinogens. Cells of the 10th, 31st, 60th and 85th passages were present in progressive development after being transfected with HPV. Cells were cultivated in a culture flask and 24-hole cultural plates. Progressive changes of morphology, cell growth, contact-inhibition, and anchorage-dependent growth characteristics were examined by phase contrast microscopy. The cell proliferation rate was assayed by flow cytometry. The modal number of chromosomes was analyzed. HPV18E(6)E(7) was detected by Western blot methods and activities of telomerase were analyzed by TRAP. Tumorigenicity of cells was detected with soft agar plates cultivated and with tumor formation in SCID mice. RESULTS: In morphological examination the 10th passage cells were in good differentiation, the 60th and 85th passages cells were in relatively poor differentiation, and the 31st passage cells had two distinct differentiations. The characteristics of the 85th and 60th passage cells were weakened at contact-inhibition and anchorage-dependent growth. Karyotypes of four stages of cells belonged to hyperdiploid or hypotriploid, and bimodal distribution of chromosomes appeared in the 31st and 60th passage cells. All of these characteristics combined with a increasing trend. The activities of telomerase were expressed in the latter three passages. Four fourths of SCID mice in the 85th passage cells and one fourth of SCID mice in the 60th passage cells developed tumors, but the cells in the 10th and 31st passage displayed no tumor formation. CONCLUSION: In continual cultivation of fetal esophageal epithelial cells with transduction of HPV18E(6)E(7), cells from the 10th to the 85th passage were changed gradually from preimmortal, immortal, precancerous to malignantly transformed stages. All of these changes were in a dynamic progressive process. The establishment of a continuous line of esophageal epithelium may provide a in vitro model of carcinogenesis induced by HPV.

Aneuploidy↗