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Biomedical subjects

J Cheng

Publications and source records attributed to J Cheng.

At least 91 records · Page 5Linked to original sources
PubMed2001

Purification and functional reconstitution of intact ral-binding Gtpase activating protein, RLIP76, in artificial liposomes.

We have recently shown that RLIP76, a ral-binding GTPase activating protein, mediates ATP-dependent transport of glutathione-conjugates (GS-E) and doxorubicin (DOX) (S. Awasthi et al., Biochemistry 39,9327,2000). Transport function of RLIP76 was found to be intact despite considerable proteolytic fragmentation in preparations used for those studies, suggesting either that the residual intact RLIP76 was responsible for transport activity, or that the transport activity could be reconstituted by fragments of RLIP76. If the former were true, intact RLIP76 would have a much higher specific activity for ATP-hydrolysis than the fragmented protein. We have addressed this question by comparing transport properties of recombinant RLIP76 and human erythrocyte membrane RLIP76 purified in buffers treated with either 100 or 500 microM serine protease inhibitor, PMSF. The purity and identity of recombinant and human erythrocyte RLIP76 was established by SDS/PAGE and Western-blot analysis. These studies confirmed the origin of the 38 kDa protein, previously referred to as DNP-SG ATPase, from RLIP76. Higher PMSF concentration resulted in lower yield of the 38 kDa band and higher yield of intact RLIP76 from both human and recombinant source. In contrast, the substrate-stimulated ATPase activity in presence of DNP-SG, doxorubicin, daunorubicin, or colchicine were unaffected by increased PMSF; similarly, ATP-dependent transport of doxorubicin in proteoliposomes reconstituted with RLIP76 was unaffected by higher PMSF. These results indicated that limited proteolysis by serine proteases does not abrogate the transport function of RLIP76. Comparison of transport kinetics for daunorubicin between recombinant vs human erythrocyte RLIP76 revealed higher specific activity of transport for tissue purified RLIP76, indicating that additional factors present in tissue purified RLIP76 can modulate its transport activity.

ATP-Binding Cassette Transporters↗
PubMed2001

[Identification of key toxicants in a bleaching effluent--a case study].

The toxicity identification evaluation (TIE) was conducted with a toxic bleaching effluent to identify its toxic components. Toxicity characterisation procedures (phase I) indicated that the primary toxicants in the effluent were volatile, and could be reduced by sodium thiosulfate. Toxicity-based fractionation techniques (phase II) were conducted, and the oxidation organic chloridates were found not to be the major toxicants. Deletion approach of phase III confirmed that the toxicity decreased with the decrease of free and total residual chlorine concentrations, indicating that Cl2, HClO and ClO- were identified as primary toxicants in the effluent.

Chlorine↗
PubMed2001

ATP sensitive K+ channel may be involved in the protective effects of preconditioning in isolated guinea pig cardiomyocytes.

OBJECTIVE: To develop a cellular model of preconditioning by a brief period of hypoxia in isolated guinea pig cardiomyocytes and to determine whether or not an ATP sensitive K+ (KATP) channel is involved in ischemic preconditioning. METHODS: Single myocytes were isolated from the ventricle of adult guinea pigs. The experimental chamber allowed the cells to be exposed to low O2 pressure. During hypoxic preconditioning, the cells were equilibrated with normaxic solution for 10 minutes and then exposed to hypoxia for 5 minutes, followed by 10 minutes of reoxygenation. The cells were then subjected to 20-180 minutes of hypoxia and reoxygenation. Ionic currents were studied with the patch clamp technique in whole-cell and cell-attached configurations. RESULTS: A 5-minute hypoxic preconditioning offered a significant protection from cell injury in subsequent hypoxia-reoxygenation. After a latency of more than 15 minutes, hypoxia induced a time-independent outward K+ current which could be blocked by 5 mumol/L glibenclamide. At 10 mV, the current increased from 78 +/- 15 pA to 1581 +/- 153 pA (P < 0.01, n = 18). However, the latency to develop KATP channel currents (IKATP) was greatly shortened in preconditioned cells, and the current was increased acceleratively. At 10 mV, the current more than 4 nA was recorded in preconditioning cells. In the single channel recordings, the time interval from the first channel opening to maximum opening was also markedly abbreviated in preconditioned cells. CONCLUSION: Isolated guinea pig cardiomyocytes can be preconditioned with a brief period of hypoxia. This hypoxic preconditioning may modify the KATP channel, and make the channel open more readily during the second hypoxia.

Adenosine Triphosphate↗
PubMed2001

bcl 10 gene mutation in hepatocellular carcinoma.

OBJECTIVE: To detect the mutation frequency of the bcl 10 gene in the early and advanced stages of hepatocellular carcinoma (HCC). METHODS: Genome DNA samples were extracted from 46 cases of fresh HCC tumor tissues and their non-tumor adjacent tissues. Polymerase chain reaction-single strand conformation polymorphism method was used to detect point mutations of the three exons of the bcl 10 gene. For each individual exon, six random samples from those showing abnormal DNA bands were sequenced to verify those mutations. The relationship between serum alpha-fetoprotein (AFP) level and bcl 10 mutation, between the tumor size and bcl 10 mutation was also analyzed. RESULTS: Among the 46 samples, 26 cases (56.5%) were found to have mutations in exon 1, 5 out of the 6 cases were shown to have 5744 C-->G mutation by sequencing; 25 cases (54.3%) were found to have mutations in exon 2, 4 out of the 6 cases were shown to have 11,311 T deletion mutation by sequencing. Twenty-one cases (45.7%) were found to have mutations in exon 3, all of the 6 cases selected for sequencing were shown to have 14,116 C-->T mutation. Statistical analysis showed that neither serum alpha-fetoprotein level nor the size of hepatocellular carcinoma has a significant relationship with bcl 10 mutation. CONCLUSION: The bcl 10 gene has a high mutation frequency in liver cancer.

Adaptor Proteins, Signal Transducing↗
PubMed2001

[Biocompatibility of HA/TCP biphasic ceramics with co-cultured human osteoblasts in vitro].

The biocompatibility of HA/TCP ceramic was evaluated by investigation of attachment and growth of osteoblasts on biomaterial, as well as monitoring the effects of biomaterial on expression of functional phenotypes of co-cultured osteoblasts in vitro. When co-cultured with HA/TCP ceramics, osteoblasts firstly attached to the surface of HA/TCP disk, then attached to notches and grew into the micropores of biomaterial during further culture period. At last, the ceramics were almost packed with osteoblasts. Additionally, osteoblasts co-cultured with HA/TCP were similar to osteoblasts cultured under normal condition in osteoblastic phenotypes; the secreted lots of collagen type I, possess strong activity of Alkaline Phosphatase and mineralized extracellular matrix. The fact that osteoblasts could grow well on HA/TCP ceramics and the biomaterial did not affect their physiological function suggest that HA/TCP ceramic is biocompatible with human osteoblasts.

Biocompatible Materials↗
PubMed2001

[Effects of air staging with absorbents on trace metal during coal combustion].

Staged combustion was carried out on laboratory-scale pulverized coal combustion with different absorbents. The experiment indicated staged combustion increased emission of submicron particles, which went against the control of trace elements, especially for those of high volatile elements, such as Cu, Ni. The thermodynamics calculation also indicate the transformation of trace metal was different with different atmosphere, suboxidized and reduced species were more easily formed under reduced condition. In both conditions, absorbents show a certain absorptive ability to trace metal, and different absorbent had different ability. For unstaged combustion, kaolinite was the best for Co, Cr and Ni; dolomite for Be, and CaO for Cu. But for under staged condition, HZ- dolomite was the best for Be, Cr and Ni; Kaolinite for Co and Cu.

Absorption↗
PubMed2001

[The role of released cytochrome C from mitochondria in the apoptosis of HUVECs induced by hydrogen peroxide].

OBJECTIVE: To explore the role of mitochondria and from which released cytochrome C in the apoptosis of HUVECs induced by H2O2. METHODS: HUVECs were cultured with H2O2 of 100 mol/L for 24 hrs. H2O2 was added after that HUVECs were precultured with CsA (cyclosporin A) for 30 mins. The cell samples were collected at different time points for DAPI staining and counting of apoptotic cell number. Simultaneously, the changes of mitochondrial permeability was observed by Rhoadmine 123 accumulation. The changes of the cytochrome C concentrations in plasma and mitochondria were determined by western blot. RESULTS: HUVECs exhibited obvious apoptosis after being processed by H2O2. The apoptotic cell number increased since 4 hrs of culturation of HUVECs with H2O2, and reached peak level at 12 hrs. And the HUVEC apoptosis induced by H2O2 could be inhibited significantly by CsA. H2O2 could lead to the decrease of mitochondrial cytochrome C concentration, which in turn lead to the increase of cytoplasmic cytochrome C concentration. Similarly, mitochondrial Rhoadmine 123 concentration decreased and the mitochondrial permeability increased. But CsA could obviously inhibit the changes of both mitochondrial Rhoadmine 123 and permeability. CONCLUSION: H2O2 could induce cytochrome C releasing to cytoplasma which lead to endothelial apoptosis. And CsA could inhibit the apoptosis by maintaining the normal function of mitochondrial membrane.

Apoptosis↗
PubMed2001

[Epidemiological observation on effect of Leptospiral outer membrane vaccine].

OBJECTIVE: To study the safety and effect of Leptospiral outer membrane vaccine. METHODS: Eighty thousands dosages of Leptospiral outer membrane vaccine were vaccinated in Jingzhou and Shishou city Hubei prevince. Temperaure, side-effects such as Local edema with in 48 hours as well as the incidence of Leptospirsis within a year among those were vaccinated and unvaccinated were observed. RESULTS: (1) No any severe side-effect and abnormal reaction, was found, only 2 case suffered from slight fever and local edema which receded in 48 hrs. (2) Effects of Leptospiral outer membrane vaccine were as follows: 2 cases were attacked by lacterohaemorrhagiae in vaccination group and 47 cases in control group, so the protection rate 95.57% and confidence interval (CI) was 85.43% - 98.20%. Fifteen hebdomadis cases were found in control group. The protection rate of this vaccine reached 100.00%, CI 77.08% - 100.00%. CONCLUSION: Safty and protective effect were well showed when type of bacteria was concordant with that in vaccination district.

Adolescent↗
PubMed2001

[The effect of Lipopolysacharide (LPS) on morphology and function of human umbilical endothelial cells (HUVECs)].

OBJECTIVE: To study the effects of LPS on the morphology and function of HUVECs, so as to explore the roles of activated vascular endothelial cells (VEC) by LPS in systemic inflammatory reaction, sepsis and MOF. METHODS: HUVECs primarily cultured in vitro were employed as the model. Inverted microscope was used to observe the effects of LPS on the morphology of HUVECs. ELISA was used to assess the IL-6 content. The expression of ICAM-1 was determined by immunofluorescent staining method with confocal laser-scanning fluorescence microscope. RESULTS: The morphology of HUVECs could be altered by LPS to become long cord-like in shape. The smallest dose of LPS for stimulating the secretion of IL-6 from HUVEC was 1 ng/ml, and the effect was enhanced parallel to the increase of the concentration of LPS and reached top level at 8 postburn hours. The expresson of ICAM-1 on the nucleic and cytoplasmic membrane of HUVECs increased obviously after HUVECs was cultured with LPS (10 ng/ml) for 24 hours when observed by laser confocal scanning light microscope. CONCLUSION: The morphology and function of HUVECs could be evidently affected by LPS. As a result, LPS might play important roles in the increase of vascular permeability, the promotion of leukocytic adherence, and the initiation of inflammatory cascade reaction.

Cell Line↗
PubMed2001

[Screening and characterization of human phage antibody to hepatitis virus C NS5A antigen].

OBJECTIVE: To screen and characterize human phage antibody against hepatitis C virus (HCV)NS5A antigen. METHODS: The recombinant phages were panned by recombinant HCV NS5A antigen which was coated in a microtiter plate; after five rounds of biopanning, 35 clones were obtained and demonstrated specific to NS5A antigen. The specific binding capacity of the ScFv antibody to HCV NS5A antigen was determined by ELISA. RESULTS: HCV NS5A phage antibody had a specific combination capacity with hepatitis C virus NS5A antigen. The DNA sequence data showed that the ScFv gene was composed of 789 bp and codes for a peptide of 262 amino acid residues. CONCLUSIONS: Human single chain antibody to hepatitis C virus NS5A antigen has been identified by means of the phage display technology.

Antibodies, Viral↗
PubMed2001

[Histopathological observation of experimental slight viral myocarditis].

To study the diagnostic method of slight viral myocarditis in the field of forensic pathology, slight viral myocarditis model was induced in Balb/c murine by coxsackie virus B3. Organs of hearts, livers, spleens, lungs and kidneys were examined through routine pathological methods. Pathological changes at different levels of these organs were observed. The results indicated that viral myocarditis was a kind of disease with multiple organ alterations and that the pathological observation and comprehensive analysis of multiple organs was one of the useful methods for diagnosing slight viral myocarditis.

Animals↗
PubMed2001

[Ultrastructure changes of the olfactory epithelium of the patients suffering from dysosmia caused by the chronic sinusitis].

OBJECTIVE: To observe the ultrastructural changes of olfactory epithelium (OE) in patients suffering from dysosmia caused by chronic sinusitis. METHODS: The specimens of olfactory epithelium were obtained from 35 patients operated for chronic sinusitis accompanied by dysosmia. According to the results of light microscope (LM) examination, the OE was divided into three groups by the types of pathological changes: normal, atrophic and respiratory epithelium metaplasia(REM). Transmission electron microscope was used to observe the ultrastructural changes of each group. RESULTS: Under the LM, the surface ultrastructure of the OE showed some abnormal changes: (1) surface microvillus of the supporting cells disappeared; (2) olfactory vesicle changed their shape due to vacuolization; (3) disappearance of canaliculus structure in the olfactory vesicle; (4) the olfactory cilia changed the shape or reduced; some of the reduced cilia underwent metaplasia. The ultrastructural changes of atrophic OE included: (1) minor and moderate atrophy: the organelles and the membrane-limited electron dense vesicles on the upper section of the supporting cells obviously decreased or disappeared, even underwent vacuolization. The basic cell degenerated; (2) serious atrophy: the turbidity of the cell structure, even double cell structure, the nuclei of the cell aggregated as the plaque and vesiculose change or karyopyknosis. As for the cytoplasm, there were the dilation of the ERs, turgidity of the mitochondrion, the disarrangement, diminution and vacuolization. Fasciculate cilia were distributed separately in the REM group. CONCLUSION: There is a positive relationship between the atrophy degree and the degree of the abnormal ultrastructural changes of the OE. The ultrastructural changes of OE in patients suffering from dysosmia caused by chronic sinusitis may provide reference for assessment of the treatment of dysosmia.

Adult↗
PubMed2001

Effects of the HOE 694 on transient inward current and Na(+) - Ca(2+) exchange in guinea pig cardiomyocytes.

OBJECTIVE: To determine the effects of HOE 694, a new and potent Na(+)-H+ exchanger blocker, on transient inward current (It(i)) and Na(+) - Ca(2+) exchange during hypoxia-reoxygenation in guinea pig cardiomyocytes. METHODS. Cardiomyocytes were isolated from adult guinea pig ventricle. Experiment was performed in an experimental chamber that allowed the cells to be exposed to a sufficiently low O2 pressure. The cells were subjected to hypoxia and reoxygenation. The ionic currents were studied with patch clamp technique. RESULTS: In the absence of HOE 694, hypoxia-reoxygenation induced It(i) in 12 of 15 experiments; but in cardiomyocytes pretreated with HOE 694 (10 approximately 50 micro mol/L), the incidence of It(i) observed during reoxygenation was reduced to 5 of 11 experiments and 3 of 10 experiments, P < 0.05 vs control respectively. The Na(+) - Ca(2+) exchange current was unaffected by HOE 694 under normoxic condition. However, when cells were pretreated with 10 micromol/L HOE 694 for 10 min, then subjected to hypoxia condition, the Na(+) - Ca(2+) exchange current significantly inhibited. CONCLUSIONS: Blockade of the Na(+)-H+ exchange by HOE 694 could reduce Ca(2+) overload upon hypoxia-reoxygenation, and inhibition of Na(+) - H+ exchange may also indirectly decrease Na(+) - Ca(2+) exchange activity during hypoxia.

Animals↗
PubMed2000

New technique for increasing retention of arginine on an anion-exchange column.

A method is described for enhancing retention of arginine on a pellicular anion-exchange column. Arginine exhibits adjustable increases of retention time that are dependent on the acidity of standard or sample matrix. This effect is based on interactions of the protonated form of arginine with the residual cation-exchange groups on the core beads of pellicular particles. The relative magnitude of retention time shift of arginine is evaluated for identical concentrations of hydrochloric, sulfuric, and perchloric acids. Although the direct addition of acid is very effective in influencing the retention of arginine, it affects peak shapes and retention of other peaks in the chromatographic separation. The new technique-acid coinjection-achieves a similar retention enhancement for arginine with only a minimal effect on the rest of the separation. Detection limits, reproducibility results, and calibration data are presented for the chromatography of amino acids with acid coinjection. Improved resolution of arginine is demonstrated with chromatograms of soybean hydrolysate and cell culture samples.

Arginine↗
PubMed2000

Complementary DNA structure and genomic organization of Drosophila menin.

Menin is a protein product of a tumor supressor gene MEN1, mutations of which are responsible for multiple endocrine neoplasia type 1, an autosomal dominant familial cancer syndrome. We determined the nucleotide sequence of the Drosophila menin cDNA using RT-PCR and RACE, and confirmed it by direct sequencing of genomic DNA. Gene expression of Drosophila menin was detected by Northern blot analysis in adult and embryo as two types of transcripts, one identical in size to the cDNA, and the other larger but detected only in embryo. The Drosophila menin gene was composed of five exons in which the protein was encoded in exon 2 through 5, and spanned approximately 6.3 kb. The deduced amino acid (AA) sequence of Drosophila menin consisted of 751 AAs with a calculated molecular mass of 81.7 kDa, and showed 44-47% identity to human, rat, mouse and zebrafish menin over the entire length. Among the AA residue substitutions that have been reported as disease-associated missense mutations and single AA deletions, 53 out of 71 were completely conserved in Drosophila. The presence of menin ortholog in insect indicates that menin is an evolutionally conserved protein with a fundamental role in biological processes.

Amino Acid Sequence↗
PubMed2000

Decrease of neuronal nitric oxide synthase in the cerebellum of aged rats.

Nitric oxide (NO) is produced as an important neurotransmitter in the central nervous system (CNS) to participate in some pathophysiological pathways. In the present study, change of neuronal nitric oxide synthase (nNOS) was examined in isolated cerebellum of Wistar rats aged from 2 to 24 months. Northern blot showed a lower mRNA level of nNOS in rats aged 6, 12 and 24 months than that in rats aged 2 months. Western blot analysis also indicated that the expression of nNOS protein was lower in rats aged 6, 12 and 24 months than that of 2 months rats. However, the activity of nNOS determined by conversion of [(3)H] L-arginine to [(3)H] L-citrulline was decreased significantly in rats aged 24 months only. These results indicate the decrease of NOS expression in cerebellum of aged rat that seems helpful to explain the causes of malfunction in CNS of aged mammalian.

Aging↗
PubMed2000

A target within the target: probing cruzain's P1' site to define structural determinants for the Chagas' disease protease.

BACKGROUND: Cysteine proteases of the papain superfamily are present in nearly all groups of eukaryotes and play vital roles in a wide range of biological processes and diseases, including antigen and hormone processing, bacterial infection, arthritis, osteoporosis, Alzheimer's disease and cancer-cell invasion. Because they are critical to the life-cycle progression of many pathogenic protozoa, they represent potential targets for selective inhibitors. Chagas' disease, the leading cause of death due to heart disease in Latin American countries, is transmitted by Trypanosoma cruzi. Cruzain is the major cysteine protease of T cruzi and has been the target of extensive structure-based drug design. RESULTS: High-resolution crystal structures of cruzain bound to a series of potent phenyl-containing vinyl-sulfone, sulfonate and sulfonamide inhibitors have been determined. The structures show a consistent mode of interaction for this family of inhibitors based on a covalent Michael addition formed at the enzyme's active-site cysteine, hydrophobic interactions in the S2 substrate-binding pocket and a strong constellation of hydrogen bonding in the S1' region. CONCLUSIONS: The series of vinyl-sulfone-based inhibitors examined in complex with cruzain was designed to probe recognition and binding potential of an aromatic-rich region of the enzyme. Analysis of the interactions formed shows that aromatic interactions play a less significant role, whereas the strength and importance of hydrogen bonding in the conformation adopted by the inhibitor upon binding to the enzyme was highlighted. A derivative of one inhibitor examined is currently under development as a therapeutic agent against Chagas' disease.

Animals↗