Biomedical subjects

M Sun

Publications and source records attributed to M Sun.

At least 163 records · Page 9Linked to original sources

PubMed1998

[Synthesis of hirudin variant 1 (HV1) gene and primary study of expression in yeast].

OBJECTIVE: Hirudin is an extremely efficient and specific thrombin inhibitor. It is clinically used to prevent the formation of thrombus. In this research the hirudin gene was put into yeast system for expression to evaluate the feasibility of artificially synthesized gene expressed in eukaryotic system and study the factor affecting expression level. METHODS: According to the amino acid sequence of hirudin variant 1 (HV1), the genetic code saccharomyces cerevisiae was used to design and synthesize the HV1 gene. Amplified by PCR, it was inserted into cloning vector pBS-SK(+) and sequenced. Ligation with the signal peptide gene of yeast alpha factor the correct HV1 gene was inserted into yeast expression vector pYC-DE. The recombinant plasmid was transformed into the cell of S. cerevisiae BJ1990 to carry out the primary expression experiment. RESULTS: In cultured supernatant of screened positive clone the hirudin activity was detected to be 30 ATU/ml. The expression level was higher than HV2 in yeast and HV1 in prokaryotic system. The N terminal amino acid sequence completely matches with natural hirudin. CONCLUSIONS: It was proved by this study that the synthesized hirudin gene had been expressed in yeast successfully. This result showed that it was a better way to carry out the expression in yeast using synthesized HV1 gene and a stronger promoter.

Base Sequence↗

PubMed1998

[Expression characteristic of Bacillus thuringiensis cry1 gene in Pseudomonas fluorescens Pfx-18].

The plasmids of lepidopteran-specific Bacillus thuringiensis strain from our laboratory were hybridized with RNA probe of cry 1Aa EcoR I-F fragment labelled using DIG. Cry1 gene was located in 39.3 MD plasmid. The plasmid was digested with Hind III and analysed by southern blot. It appeared both 7.1 kb and of 6.5 kb positive bands. The 7.1 kb fragment was ligated to broad-host-range vector pSUP106 and transformed into Pseudomonas fluorescens Pfx-18. The cloned strain, LZP-1 was obtained. The plasmids of LZP-1 were analysed by PCR. The results showed that gene-type is cry1Ab. SDS-PAGE analysis demonstrated that LZP-1 could express 66 kD insecticidal crystal protein and some small molecular weight peptides. Bioassay showed that motality of 1000-fold diluted fermentation broth was 33% to 3rd instar plulella xyloslelly larvae.

Animals↗

PubMed1998

Deposition of advanced glycation end products (AGE) and expression of the receptor for AGE in cardiovascular tissue of the diabetic rat.

Advanced glycation end products (AGE) in tissues are important for the central pathological features of diabetic complication. Although AGE bind to several cell-surface sites, resulting in altered cellular functions, receptor for AGE (RAGE) appears to have a central role. We examined AGE accumulation and RAGE expression in the aorta and heart of rats with streptozotocin (STZ)-induced diabetes, 0, 4, 8, 12, 16 and 24 weeks after STZ administration. Early atherosclerotic findings in the intima and medial thinning were observed in the aorta after 16 weeks of STZ-Induced diabetes. Immunohistochemistry and microscope spectrophotometry showed that AGE deposition increased significantly in the aorta and vessels of the myocardium, depending on the period of hyperglycaemia. RAGE was expressed in the endothelial cells and vascular smooth muscle cells of all animals. The number of smooth muscle cells with RAGE immunoreactivity increased until 12 weeks after STZ injection, and then decreased in rats with diabetes between 16 and 24 weeks. On the other hand, total RAGE mRNA levels in the aorta and heart continued to increase with the duration of hyperglycaemia. Furthermore, AGE-BSA induced RAGE mRNA expression of human umbilical vein endothelial cells in vitro. Taken together, the AGE accumulation might initiate diabetic macroangiopathy through RAGE, and the increase of RAGE expression by endothelial cells could be a reason that diabetes mellitus accelerates atherosclerosis rapidly.

Animals↗

PubMed1997

Formation and cleavage of a DNA network during in vitro bacteriophage T7 DNA packaging: light microscopy of DNA metabolism.

To understand in vivo DNA metabolism, in vitro systems are developed that perform DNA metabolism, while maintaining in vivo (physiological) character. To determine the state of DNA during in vitro physiological metabolism, the present study develops procedures of fluorescence light microscopy for observation of stained DNA molecules during in vitro physiological metabolism in a crude extract of bacteriophage T7-infected cells. The extract inhibits illumination-induced breakage of DNA. The following DNA metabolism remains active for 2-3 min during microscopy: exonuclease-dependent end-to-end joining (concatemerization) of T7 DNA and subsequent cleavage of concatemers. When the T7 gene 3-encoded DNA debranching endonuclease is absent during in vitro T7 DNA concatemerization, DNA progressively partitions to form a continuous, mostly immobile (i.e., no detected Brownian motion) fibrous network that encloses the DNA-depleted solution; presumably because of reduced branching, a less extensive network forms when the gene 3-encoded debranching endonuclease is present. Most strands of the network consist of multiple DNA segments. After a time interval of 5-10 min, the DNA network undergoes cleavage that depends on the presence of both ATP, capsids, and the DNA packaging accessory proteins encoded by genes 18 and 19; multiple cleavages eventually disrupt the continuity of the DNA network. The dependence of the observed cleavage on these factors is explained by the hypothesis that this cleavage is the first of two cleavages known to occur during the packaging of T7 DNA concatemers both in vivo and in vitro. The first cleavage is also known to initiate entry of DNA into a T7 capsid. The cleavage observed here is usually preceded by an approximately 10 s burst of oscillatory motion of the DNA network near the point of eventual cleavage. If the in vivo presence of a similar concatemer-containing DNA network is assumed, requirement for DNA packaging-associated release of DNA from this network is a possible explanation for the evolution of a T7 DNA packaging pathway that is initiated by cleavage of a concatemer.

Adenosine Triphosphate↗

PubMed1997

The role of dietary factors in the intestinal and diffuse histologic subtypes of gastric adenocarcinoma: a case-control study in the U.S.

BACKGROUND: The decline of gastric adenocarcinoma in the U. S. and the parallel decrease in the predominance of the intestinal type of gastric adenocarcinoma may reflect, in part, changes in diet over the last 60 years. Because the intestinal and diffuse types of gastric adenocarcinoma may be epidemiologically distinct, the authors hypothesized that different nutritional factors are associated with the different subtypes of gastric adenocarcinoma. METHODS: Ninety-one incident cases with a pathologic diagnosis of gastric adenocarcinoma and 132 cancer free controls were included in this study. All cases were defined as being either the intestinal or diffuse type of gastric adenocarcinoma. Epidemiologic data were collected by a modified National Cancer Institute Health Habits History Questionnaire. Nutritional and dietary factors were analyzed using a logistic regression model. RESULTS: Several dietary factors were significantly associated with both subtypes of gastric adenocarcinoma, including dietary intakes of fiber, oleic acid, potassium, and fruits. Almost all dietary factors and food groups unique to intestinal gastric adenocarcinoma were protective in nature (vitamin B6, folate, niacin, iron, noncitrus fruits, and raw fruit), except for a high intake of dietary calories, which was a risk factor for intestinal gastric adenocarcinoma. The unique factors found to be protective for diffuse disease were carbohydrate and vitamin C intake. CONCLUSIONS: This study suggests that dietary factors contribute to the carcinogenesis of gastric adenocarcinoma. It also appears that although dietary risk and protective factors are common to both the intestinal and diffuse types of this malignancy, protective dietary factors may play a more important role in preventing the intestinal type of gastric adenocarcinoma.

Adenocarcinoma↗

PubMed1997

Cleavage specificity of a proteolytic antibody light chain and effects of the heavy chain variable domain.

The recombinant light chain (L chain) of an antibody raised by immunization with vasoactive intestinal polypeptide (VIP) cleaved this peptide on the C-terminal side of basic residues. The major sites of cleavage in VIP were two adjacent peptide bonds, Lys20-Lys21 and Lys21-Tyr22. Lower levels of cleavage were evident at Arg14-Lys15 and Lys15-Gln16. Hydrolysis of radiolabeled VIP by the L chain was inhibited by two serine protease inhibitors, diisopropylfluorophosphate and aprotinin, but not by soybean or lima bean trypsin inhibitors or inhibitors of other classes of proteases. To probe the role of the VH domain, single chain Fv constructs composed of the VL domain of the anti-VIP L chain linked via a 14-residue peptide to its natural VH domain partner or an irrelevant anti-lysozyme VH domain (hybrid Fv) were prepared. The anti-VIP Fv hydrolyzed VIP with Ks 21.4-fold lower than the L chain and 250-fold lower than the hybrid Fv, suggesting increased affinity for the substrate ground state due to the anti-VIP VH domain. The kinetic efficiency (kcat/Ks) of the anti-VIP Fv was 6.6-fold greater compared to the L chain and 29.4-fold greater compared to the hybrid Fv. Peptide-MCA substrates unrelated in sequence to VIP were hydrolyzed by the anti-VIP Fv and L chain at equivalent rates. These observations lead to a model of catalysis by the anti-VIP Fv in which the essential catalytic residues are located in the VL domain and additional residues from the VH domain are involved in high affinity binding of the substrate.

Amino Acid Sequence↗

PubMed1997

Snmp-1, a novel membrane protein of olfactory neurons of the silk moth Antheraea polyphemus with homology to the CD36 family of membrane proteins.

While olfactory neurons of silk moths are well known for their exquisite sensitivity to sex pheromone odorants, molecular mechanisms underlying this sensitivity are poorly understood. In searching for proteins that might support olfactory mechanisms, we characterized the protein profile of olfactory neuron receptor membranes of the wild silk moth Antheraea polyphemus. We have purified and cloned a prominent 67-kDa protein which we have named Snmp-1 (sensory neuron membrane protein-1). Northern blot analysis suggests that Snmp-1 is uniquely expressed in antennal tissue; in situ hybridization and immunocytochemical analyses show that Snmp-1 is expressed in olfactory neurons and that the protein is localized to the cilia, dendrites, and somata but not the axons. Snmp-1 mRNA expression increases significantly 1-2 days before the end of adult development, coincident with the functional maturation of the olfactory system. Sequence analysis suggests Snmp-1 is homologous with the CD36 protein family, a phylogenetically diverse family of receptor-like membrane proteins. CD36 family proteins are characterized as having two transmembrane domains and interacting with proteinaceous ligands; Snmp-1 is the first member of this family identified in nervous tissue. These findings argue that Snmp-1 has an important role in olfaction; possible roles of Snmp-1 in odorant detection are discussed.

Amino Acid Sequence↗

PubMed1997

Altered cleavage site preference of a proteolytic antibody light chain induced by denaturation.

A recombinant antibody light chain (L chain) maintained under non-denaturing conditions displayed preferential cleavage of synthetic peptides conjugated to methylcoumarinamide (MCA) on the C-terminal side of Arg and Lys residues. The same L chain renatured from a denaturing solvent (guanidine hydrochloride) acquired the capability of cleaving Tyr-MCA and Leu-MCA bonds, and its ability to cleave MCA linked to basic residues was decreased. The altered cleavage preference was accompanied by a conformational transition in the protein, evident from the fluorescence emission spectra. These observations suggest the feasibility of redirecting the cleavage specificity via alterations in the conformation of proteolytic antibody combining sites.

Amino Acid Sequence↗

PubMed1997

The conformation of DNA packaged in bacteriophage G.

When packaged in a bacteriophage capsid, double-stranded DNA occupies a cavity whose volume is roughly twice the volume of the DNA double helix. The data thus far have not revealed whether the compactness of packaged bacteriophage DNA is achieved by folding of the DNA, undirectional winding of the DNA, or a combination of both folding and winding. To assist in discriminating among these possibilities, the present study uses electron microscopy, together with ultraviolet light-induced DNA-DNA cross-linking, to obtain the following information about the conformation of DNA packaged in the comparatively large bacteriophage, G: 1) At the periphery of some negatively stained particles of bacteriophage G, electron microscopy reveals standards of DNA that are both parallel to each other and parallel to the polyhedral bacteriophage G capsid. However, these strands are not visible toward the center of the zone of packaged DNA. 2) Within some positively stained particles, electron microscopy reveals DNA-associated stain in relatively high concentration at corners of the polyhedral bacteriophage G capsid. 3) When cross-linked DNA is expelled from its capsid during preparation for electron microscopy, some DNA molecules consist primarily of a compacted central region, surrounded by DNA strands that appear to be unravelling at multiple positions uniformly distributed around the compacted DNA region. The above results are explained by a previously presented model in which DNA is compacted by folding to form 12 icosahedrally arranged pear-shaped rings.

Bacteriophages↗

PubMed1997

ICV beta-hydroxybutyrate: effects on food intake, body composition, and body weight in rats.

This study examined the effect of long-term intracerebroventricular (ICV) infusion of beta-hydroxybutyrate (beta HB) on food intake, diet selection, body weight, and body composition in rats. Female rats were divided into 2 groups and implanted with a 28-day osmotic pump connected to a lateral cerebroventricular cannula. One group was infused with artificial cerebrospinal fluid (aCSF) and the other with beta HB for 28 days. The rats had free access to both a high-fat/low-carbohydrate and a low-fat/high-carbohydrate diet (isocaloric) for the 28-day infusion period. The group infused with beta HB had a significantly lower body weight gain during the infusion period. Cumulative food intake increased in the same manner in both groups. Fat pad weights and carcass lipid content were significantly higher in beta HB rats, despite the equivalent caloric intake in both groups and the decreased body weight. Our observations are in accord with earlier studies indicating that beta HB infused ICV reduces body weight, but not necessarily food intake. Increased adiposity in association with decreased body weight change in beta HB-infused rats strongly suggests that energy is being partitioned to fat deposition at the expense of lean tissue growth when ketone bodies are infused into the cerebroventricles.

3-Hydroxybutyric Acid↗

PubMed1997

Prognostic significance of detection of prostate-specific antigen transcripts in the peripheral blood of patients with metastatic androgen-independent prostatic carcinoma.

OBJECTIVES: To evaluate the prognostic significance of reverse transcriptase polymerase chain reaction (RT PCR) detection of prostate-specific antigen (PSA) mRNA in relation to survival in patients with metastatic androgen-independent prostatic carcinoma (AIPC). METHODS: Peripheral blood from 122 men (64 from Memorial Sloan-Kettering Cancer Center [MSKCC] and 58 from the Dana Farber Cancer Institute [DFCI]) with metastatic (Stage D2) AIPC was analyzed for PSA mRNA using RT PCR. Forty-one controls without prostatic carcinoma were also evaluated. RESULTS: RT PCR positivity for PSA mRNA was present in 24 of the 64 (38%) patients seen at MSKCC and in 26 of the 58 (45%) patients followed at DFCI. All control individuals were PSA PCR negative. There was a significant correlation between RT PCR positivity and decreased survival in each of the Memorial and Dana Farber population (P = 0.028 and 0.039, respectively). Serum PSA (at time of blood collection for PCR) was not predictive of survival as a continuous variable in the MSKCC [P = 0.31] and the DFCI (P = 0.09) groups. RT PCR for PSA mRNA was found to be independent from and superior to serum PSA in predicting survival in both the MSKCC and DFCI populations (P = 0.048 and P = 0.027, respectively). CONCLUSIONS: The detection of PSA mRNA in the peripheral blood by RT PCR is a predictor of survival in patients with metastatic AIPC, and PCR is superior to a single serum PSA measurement. Further studies are needed to test the value of this factor in comparison to and coupled with other prognostic parameters.

Adult↗

PubMed1997

Cocaine and inhibition of nitric oxide synthesis produce opioid-mediated antinociception.

Cocaine and nitric oxide are known to influence the perception of pain. The present study sought to determine if the endogenous opioid peptide system participates in cocaine-induced antinociception and antinociception produced by antagonism of nitric oxide. Pain perception was measured using the hot plate test. Administration of cocaine (25 mg/kg) to male rats resulted in a significant increase in reaction time in the hot plate test, which was reversed by treatment with 3, 10, and 30 mg/kg of the opiate antagonist, naloxone. In rats that were not treated with cocaine, doses of 30 and 60 mg/kg of naloxone significantly reduced hot plate reaction Time. Treatment of animals with the nitric oxide synthase inhibitor, N omega nitro-L-arginine, produced a significant increase in response time to the hot plate, which was reversed by administration of naloxone. These data indicate that antinociception produced in the rat by cocaine appears to have a supraspinal component and to involve activation of endogenous opioid peptide activity in the brain. The results also suggest a tonic inhibition of endogenous opioid peptide activity by nitric oxide, which when antagonized, results in diminished response to pain.

Animals↗

PubMed1997

Adenocarcinomas of the esophagus and gastric cardia: the role of diet.

The incidence of adenocarcinomas of the esophagus and gastric cardia (ACEGC) has been increasing for the past 10-15 years in the United States. The reason for this increase is unknown. This hospital-based case-control study was conducted to assess the effects of dietary and nutritional factors on the risk of ACECG. A total of 95 incident cases with pathological diagnosis and 132 cancer-free controls were included in the study. Patients were recruited at Memorial Sloan-Kettering Cancer Center from 1 November 1992 to 1 November 1994. Epidemiologic data were collected by a modified National Cancer Institute Health Habits History Questionnaire. Nutritional and dietary factors were analyzed using a logistic regression model. Increased risk of ACEGC was significantly related to higher intake of dietary calories and fat after controlling for several potential confounding factors. Decreased risk of ACEGC was significantly associated with high ingestion of dietary fiber, lutein, niacin, vitamin B6, iron, and zinc. Higher intakes of vitamin A, beta-carotene, vitamin E, folate, phosphorus, and potassium were associated with a decreased risk of the disease, but these were not statistically significant. The study suggests that ACEGC can be preventable through dietary interventions.

Adenocarcinoma↗

PubMed1997

Genetic susceptibility to head and neck cancer: interaction between nutrition and mutagen sensitivity.

The development of head and neck cancer may depend not only on exposure to environmental carcinogens but also on a genetically based susceptibility to carcinogen-induced damage. This thesis presents a case-control study that demonstrates the significance of mutagen sensitivity, a measure of an individual's intrinsic DNA repair capacity against free radical damage, as a risk factor for the disease. As part of the case-control analysis, 167 previously untreated patients and 177 age- and sex-matched healthy controls were assessed for various lifestyle factors including tobacco and alcohol habits, occupational exposures, and diet. Mutagen sensitivity expressed by each individual was determined by quantifying bleomycin-induced chromosomal breaks within peripheral blood lymphocytes in vitro. Consistent with our initial observations and those of others, mutagen hypersensitivity was strongly associated with increased risk of head and neck cancer (odds ratio, 4.95; 95% confidence interval, 2.67 to 9.17) after adjusting for age, sex, and race. Low intake of vitamins C and E was also associated with an increased risk of disease and was interactive with mutagen sensitivity in risk estimates. Individuals with both a low intake of various antioxidants and increased chromosomal sensitivity to oxidant-induced DNA damage were at greatest risk. This study supports the concept that the risk of head and neck cancer is determined by a balance of factors that either enhance or protect against free radical oxygen damage, including innate capacities for DNA repair.

Adult↗

PubMed1997

An efficient algorithm for computing multishell spherical volume conductor models in EEG dipole source localization.

Computationally localizing electrical current sources of the electroencephalographic signal requires a volume conductor model which relates theoretical scalp potentials to the dipolar source located within the modeled brain. The commonly used multishell spherical model provides this source-potential relationship using a sum of infinite series whose computation is difficult. This paper provides a closed-form approximation to this sum based on an optimal fitting to the weights of the Legendre polynomials. The second-order (third-order) approximation algorithm, implemented by a provided C-routine, requires only 100 (140) floating point operations to compute a single scalp potential in response to an arbitrary current dipole located within a four-shell spherical volume conductor model. This cost of computation represents only 6.3% (8.9%) of that required by the direct method. The relative mean square error, measured by using 20,000 random dipoles distributed within the modeled brain, in only 0.29% (0.066%).

Algorithms↗

PubMed1997

Efficacy of prophylactic aerosol amphotericin B lipid complex in a rat model of pulmonary aspergillosis.

Invasive pulmonary aspergillosis remains an important cause of morbidity and mortality among transplant recipients and patients receiving cancer chemotherapy. The lipid-associated formulation of amphotericin B (AmB), AmB lipid complex (ABLC), was evaluated for its prophylactic efficacy when it was administered as an aerosol in a rat model of pulmonary aspergillosis. Aerosol ABLC (aero-ABLC), in doses from 0.4 to 1.6 mg/kg of body weight given 2 days before infection, significantly delayed mortality compared to the mortality of rats given placebo (P < 0.001). At day 10 postinfection, 50% of rats in the 0.4-mg/kg group and 75% of rats in the 1.6-mg/kg group were alive, while all control animals had died. In a second trial aero-ABLC was more effective than an equivalent dose of aerosol AmB (aero-AmB) in prolonging survival, with 100% survival at day 14 postinfection in the ABLC group, compared to 62.5% survival in the AmB group. Mean concentrations of AmB in lungs were 3.7 times higher at day 1 (P < 0.002) and almost six times higher at day 7 (P < 0.001) after treatment with aero-ABLC than after treatment with a similar dose of aero-AmB. We conclude that aero-ABLC provided higher and more prolonged levels of the parent compound in the lungs than aero-AmB and was more effective in delaying mortality from aspergillosis in this model.

Aerosols↗

PubMed1997

Improving energy expenditure estimation by using a triaxial accelerometer.

In our study of 125 subjects (53 men and 72 women) for two 24-h periods, we validated energy expenditure (EE), estimated by a triaxial accelerometer (Tritrac-R3D), by using a whole-room indirect calorimeter under close-to-normal living conditions. The estimated EE was correlated with the measured total EE for the 2 days (r = 0. 925 and r = 0.855; P < 0.001) and in minute-by-minute EE (P < 0.01). Resting EE formulated by the Tritrac was found to be similar to the measured values [standard errors of estimation (SEE) = 0.112 W/kg; P = 0.822]. The Tritrac significantly underestimated total EE, EE for physical activities, EE of sedentary and light-intensity activities, and EE for exercise such as stepping (all P < 0.001). We developed a linear and a nonlinear model to predict EE by using the acceleration components from the Tritrac. Predicted EE was significantly improved with both models in estimating total EE, total EE for physical activities, EE in low-intensity activities, minute-by-minute averaged relative difference, and minute-by-minute SEE (all P < 0. 05). Furthermore, with our generalized models and by using subjects' physical characteristics and body acceleration, EE can be estimated with higher accuracy (averaged SEE = 0.418 W/kg) than with the Tritrac model.

Acceleration↗

PubMed1997

Spatial patterns of olfactory neurons expressing specific odor receptor genes in 48-hour-old embryos of zebrafish Danio rerio.

Olfactory neurons have a complex phenotype characterized by their expression of a specific odor receptor (OR) gene and their targeting of an equally specific locus in the olfactory bulb. In the adult fish, olfactory neurons expressing specific ORs are broadly distributed in the epithelium, intermingling with neurons expressing other OR phenotypes. This distributed adult pattern has led to the suggestion that olfactory neuron phenotype is determined by a stochastic process, independent of external positional cues. However, when the fish olfactory system is established during embryogenesis it is simple in its organization, with few olfactory neurons and an olfactory epithelium that has not yet folded into the adult morphology. It is possible that positional cues might act in the embryo to establish an initial population and pattern of olfactory neuron phenotypes and that subsequent morphogenesis and neuronal addition lead to the randomized distribution of neurons. To test this possibility, we examined the spatial patterns of olfactory neurons expressing specific OR genes in 48 h embryos, a time of relative simplicity in the developing olfactory epithelium. Three-dimensional plots of neuron distributions were made, and comparison of OR expression patterns were made between right and left epithelia, between individual animals and between different OR genes. The patterns of OR gene expression were not conserved in these comparison. Mathematical analysis of 21 epithelia for the degree of order in the distribution of olfactory neurons argued strongly that the neurons expressing given ORs are randomly distributed in the 48 h embryos. These results are consistent with those observed from adult tissue and support models suggesting that extrinsic positional cues do not have a major role in specifying olfactory neuron phenotypes.

Animals↗