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Biomedical subjects

W Chen

Publications and source records attributed to W Chen.

At least 1,081 records · Page 60Linked to original sources
PubMed1992

Suppression of human nasopharyngeal carcinoma cell growth in nude mice by the wild-type p53 gene.

Wild-type and mutant human p53 genes were transfected into the nasopharyngeal carcinoma (NPC) cell line CNE-3. Tumorigenicity in nude mice showed that the tumor resulting from the cells transfected with the wild-type p53 gene grew more slowly and was smaller than that from the cells transfected with mutant p53 gene and that from control CNE-3 cells. In contrast, the tumor from the cells transfected with the mutant p53 gene grew faster than that produced by cells transfected with the wild-type p53 gene and that produced by control CNE-3 cells. The results demonstrate that the wild-type p53 gene could inhibit the NPC cell growth in nude mice and the mutant p53 gene could enhance the NPC cell growth in nude mice. The p53 gene may also play an important role in the pathogenesis of NPC.

Animals↗
PubMed1992

Restricted replication of ectromelia virus in cell culture correlates with mutations in virus-encoded host range gene.

Ectromelia virus (strain Moscow) was shown to replicate poorly or not at all in cell lines derived from the rabbit or hamster. The failure of ectromelia virus to replicate in cell lines derived from the hamster suggested that the virus lacked a functional CHO host range (hr) gene required for multiplication in these cells. A DNA fragment which hybridized to the CHO hr gene was cloned from the ectromelia virus genome and shown by sequence analysis to be deleted of 506 bp within the ectromelia virus CHO hr homologue. Two additional ectromelia viruses (Hampstead and Mill Hill strains) were also shown to lack an intact CHO hr gene. Insertion of the CHO gene from cowpox virus into the ectromelia virus genome extended the host range of ectromelia virus in tissue culture. These results demonstrate that an intact CHO hr gene is not required for maintenance of ectromelia virus in nature and provide a partial explanation for ectromelia virus' narrow host range, as opposed to the broad host range of cowpox virus, which has a functional CHO hr gene.

Animals↗
PubMed1992

A simple and rapid method of preparing large fragments of dengue virus cDNA from replicative-form RNA using reverse transcriptase and PCR.

A method is described for cloning large fragments (1.5 kb to 2 kb) of dengue virus cDNA from replicative-form viral RNA. Aedes albopictus cells (C6/36 clone) infected with dengue virus contain double-stranded, replicative-form RNA molecules which were used as a template for an initial reverse transcription using a primer containing sequence homologous to regions of the genome at or near the 3' end of the gene being studied. The product was then used as a template for polymerase chain reaction (PCR) amplification using the same 3' primer and a second which hybridized to a region at the 5' end of the sequence to be cloned. Both primers were engineered to contain specific restriction enzyme cutting sites which enabled the PCR product to be cut and cloned directly into plasmids for sequencing and expression studies. We have used this method to construct clones of the envelope glycoprotein gene (E) and the non-structural genes 1 and 2a (NS1/2a) and 3 (NS3) of dengue type 2, Tonga 1974 strain, and E and NS1/2a from dengue type 3, H-87 strain, either as discrete genes or as constructs with long and short leader sequences, with or without anchor sequences. The method could be applied to the cloning of any gene from any flavivirus, directly from infected cell extracts, without the necessity for tedious virus purification steps.

Animals↗
PubMed1992

Polyclonal antibodies against the polypeptide and carbohydrate epitopes of recombinant human choriogonadotropin beta-subunit.

In our previous paper (Chen et al. (1991) J. Biol. Chem. 266, 4081-4087) we reported the preparation and characterization of recombinant human choriogonadotropin beta subunit (hCG beta) using the baculovirus-insect cell expression system. The rhCG beta was found to contain high mannose type N-linked carbohydrates and 3-4 serine-linked disaccharide chains. Despite the carbohydrate structural variation, the rhCG beta was similar to hCG beta in in vitro immunological and biological properties. In order to evaluate its in vivo immunological properties, rabbit antiserum against rhCG beta was produced. The antiserum was found to be almost identical to anti-hCG beta in binding to hCG beta as well as in its crossreactivity with human lutropin (hLH), hCG and human follitropin (hFSH) as indicated by radioimmunoassays using 125I-hCG beta as a tracer. Further characterization of the anti-rhCG beta antiserum revealed that there are three types of antibodies in terms of antigenic specificity present in the anti-rhCG beta antisera pool as shown by dot blot and radioimmunoassays. The carbohydrate-specific antibodies were separated by affinity chromatography using an ovalbumin-glycopeptide-Sepharose column. The antibodies held on the ovalbumin affinity adsorbent were specific for the high mannose type carbohydrates such as those present in rhCG beta, rhCG and thyroglobulin and failed to react with transferrin, alpha 1-acid glycoprotein and hCG alpha, all containing complex type carbohydrates. This was further supported by the fact that the recombinant unglycosylated hCG or periodate oxidized rhCG beta also did not show any reactivity with the carbohydrate specific antibodies. Two types of peptide epitopes seemed to be present in rhCG beta since when the flowthrough fraction from the ovalbumin-glycopeptide-affinity column was passed through the hCG beta-Sepharose column, the antibodies in the flowthrough from the latter column were specific to the unique antigenic determinants present only in the rhCG beta and not in hCG beta. The eluate from the hCG beta-Sepharose column contained the third type of antibodies, being the predominant ones, directed to the common epitopes between rhCG beta and hCG beta. The high mannose type specific antibodies are potentially useful in differentiating between the high mannose and complex type of N-linked carbohydrates present in a glycoprotein. Also, the antibody could provide an effective reagent in studying the intracellular processing of the N-linked oligosaccharides.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1992

Effects of conditioning depolarization and repetitive stimulation on Q beta and Q gamma charge components in frog cut twitch fibers.

Charge movement was measured in frog cut twitch fibers with the double Vaseline-gap technique. Steady-state inactivation of charge movement was studied by changing the holding potential from -90 mV to a level ranging from -70 to -30 mV. Q beta and Q gamma at each holding potential were separated by fitting the Q-V plot with a sum of two Boltzmann distribution functions. At -70 mV Q beta and Q gamma were inactivated to 54.0% (SEM 2.2) and 82.7% (SEM 3.0) of the amounts at -90 mV. At holding potentials greater than or equal to -60 mV, more Q gamma was inactivated than Q beta, and at -30 mV Q gamma was completely inactivated but Q beta was not. There was no holding potential at which Q beta was unaffected and Q gamma was completely inactivated. The differences between the residual fractions of Q beta and Q gamma are significant at all holding potentials (P less than 0.001-0.05). The plot of the residual fraction of Q beta or Q gamma versus holding potential can be fitted well by an inverted sigmoidal curve that is a mirror image of the activation curve of the respective charge component. The pair of curves for Q gamma correlates well with those for tension generation or Ca release obtained by other investigators. The time courses of the inactivation of Q beta and Q gamma were studied by obtaining several Q-V plots with conditioning depolarizations lasting 1-20 s and separating each Q-V plot into Q beta and Q gamma components by fitting with a sum of two Boltzmann distribution functions. The inactivation time constant of Q beta was found to be 5-10 times as large as that of Q gamma. During repetitive stimulation, prominent I gamma humps could be observed in TEST-minus-CONTROL current traces and normal Q gamma components could be separated from the Q-V plots, whether 20 or 50 mM EGTA was present in the internal solution, whether 2 or 10 stimulations were used, and whether the stimuli were separated by 400 ms or 6 s. Repetitive stimulation slowed the kinetics of the I gamma hump and could shift the Q-V curve slightly in the depolarizing direction in some cases, resulting in an apparent suppression of charge at the potentials that fall on the steep part of the Q-V curve.

Animals↗
PubMed1992

Separation of Q beta and Q gamma charge components in frog cut twitch fibers with tetracaine. Critical comparison with other methods.

Charge movement was measured in frog cut twitch fibers with the double Vaseline-gap technique. 25 microM tetracaine had very little effect on the maximum amounts of Q beta and Q gamma but slowed the kinetics of the I gamma humps in the ON segments of TEST-minus-CONTROL current traces, giving rise to biphasic transients in the difference traces. This concentration of tetracaine also shifted V gamma 3.7 (SEM 0.7) mV in the depolarizing direction, resulting in a difference Q-V plot that was bell-shaped with a peak at approximately -50 mV. 0.5-1.0 mM tetracaine suppressed the total amount of charge. The suppressed component had a sigmoidal voltage distribution with V = -56.6 (SEM 1.1) mV, k = 2.5 (SEM 0.5) mV, and qmax/cm = 9.2 (SEM 1.5) nC/microF, suggesting that the tetracaine-sensitive charge had a steep voltage dependence, a characteristic of the Q gamma component. An intermediate concentration (0.1-0.5 mM) of tetracaine shifted V gamma and partially suppressed the tetracaine-sensitive charge, resulting in a difference Q-V plot that rose to a peak and then decayed to a plateau level. Following a TEST pulse to greater than -60 mV, the slow inward current component during a post-pulse to approximately -60 mV was also tetracaine sensitive. The voltage distribution of the charge separated by tetracaine (method 1) was compared with those separated by three other existing methods: (a) the charge associated with the hump component separated by a sum of two kinetic functions from the ON segment of a TEST-minus-CONTROL current trace (method 2), (b) the steeply voltage-dependent component separated from a Q-V plot of the total charge by fitting with a sum of two Boltzmann distribution functions (method 3), and (c) the sigmoidal component separated from the Q-V plot of the final OFF charge obtained with a two-pulse protocol (method 4). The steeply voltage-dependent components separated by all four methods are consistent with each other, and are therefore concluded to be equivalent to the same Q gamma component. The shortcomings of each separation method are critically discussed. Since each method has its own advantages and disadvantages, it is recommended that, as much as possible, Q gamma should be separated by more than one method to obtain more reliable results.

Animals↗
PubMed1992

Importance of endogenous tumor necrosis factor alpha and gamma interferon in host resistance against Pneumocystis carinii infection.

C.B-17 scid/scid (SCID) mice that have acquired natural pulmonary infection with Pneumocystis carinii clear these organisms by 19 days after reconstitution with spleen cells from immunocompetent mice and therefore serve as a model for studying the pathogenesis of and immunity to P. carinii pneumonia. The present study examined the importance of endogenous tumor necrosis factor alpha (TNF-alpha) and gamma interferon (IFN-gamma) in the clearance of P. carinii by treatment of reconstituted SCID mice with anti-TNF-alpha and anti-IFN-gamma immunoglobulin G (IgG). Treatment of reconstituted mice with monospecific rabbit anti-TNF-alpha IgG almost completely inhibited the clearance of P. carinii from the lungs. In contrast, treatment with either anti-IFN-gamma antibody (polyclonal or monoclonal) or control IgG had no detectable effect on the clearance of P. carinii. The importance of endogenous TNF-alpha in the clearance of P. carinii was further supported by the finding of TNF-alpha but not IFN-gamma in lung homogenate supernatants from reconstituted SCID mice. Further study revealed that for the complete clearance of P. carinii, TNF-alpha must be present at the early stage of reconstitution, since clearance could be blocked by a single injection of anti-TNF-alpha IgG into SCID mice at day 0 but not at day 6 and/or day 12 after reconstitution. These results strongly suggest that, in reconstituted SCID mice, endogenous TNF-alpha is important in host resistance against P. carinii infection, whereas IFN-gamma appears not to play a significant role.

Analysis of Variance↗
PubMed1992

Resistance of severe combined immunodeficient mice to infection with Cryptosporidium parvum: the importance of intestinal microflora.

Cryptosporidium parvum is a protozoan parasite which colonizes intestinal epithelium, causing transient diarrheal illness in immunocompetent hosts and severe chronic disease in immunocompromised hosts. We examined the resistance of severe combined immunodeficient mice, either bearing intestinal flora or germfree, to intestinal infection with C. parvum. Infection was not readily detected in flora-bearing adult severe combined immunodeficient mice until 5 to 7 weeks following oral challenge with C. parvum. In contrast, germfree adult severe combined immunodeficient mice were heavily infected 3 weeks following challenge. These data support the hypothesis that resistance of adult mice to C. parvum infection does not require a specific immune response but can be mediated by nonspecific mechanisms associated with the presence of intestinal flora.

Animals↗
PubMed1992

The vaccinia virus B1R gene product is a serine/threonine protein kinase.

The nucleotide sequence of the vaccinia virus open reading frame B1 predicts a polypeptide with significant sequence similarity to the catalytic domain of known protein kinases. To determine whether the B1R polypeptide is a protein kinase, we have expressed it in bacteria as a fusion with glutathione S-transferase. Affinity-purified preparations of the fusion protein were found to undergo autophosphorylation and also phosphorylated the exogenous substrates casein and histone H1. Mutation of lysine 41 to glutamine within the conserved kinase catalytic domain II abrogated protein kinase activity on all three protein substrates, supporting the notion that the protein kinase activity is inherent to the B1R polypeptide. Casein and histone H1 were phosphorylated on serine and threonine residues. The B1R fusion protein was phosphorylated on a threonine residue(s) by an apparently intramolecular mechanism. The autophosphorylation reaction resulted in phosphorylation of the glutathione S-transferase portion of the fusion and not the protein kinase domain. The protein kinase activity of B1R was specific for ATP as the phosphate donor; GTP was not utilized to a detectable extent. Immunoblotting experiments with anti-B1R antiserum showed that the protein kinase is located in the virion particle. Chromatography of virion extracts resulted in separation of the B1R protein kinase from the bulk of the total protein kinase activity, indicating that multiple protein kinases are present in the virion particle and that B1R is distinct from the previously described vaccinia virus-associated protein kinase.

Adenosine Triphosphate↗
PubMed1992

The effect of IH764-3 on fibroblast proliferation and function.

The effect of IH764-3, a potent component isolated from Salvia miltiorrhiza, on the proliferation and function of cultured fibroblasts was studied. It was found that the fibroblast growth curve had a dose-dependent relationship with IH764-3 concentration. The incorporation of 3H-TdR and 3H-proline into fibroblasts was significantly inhibited by IH764-3, and calmodulin, fibronectin and thrombospondin contents in the test group were obviously lower than those in the control group. Flow cytometry showed that in the IH764-3-treated group, the percentage of cells in G0/G1 phase was higher than that in the control. Electron microscopic observation (TEM and SEM) showed that in the treated group, collagen secretion was decreased. All of these results indicate that IH764-3 exerts a direct inhibitory effect on fibroblast proliferation and affects their ability to synthesize collagen.

Animals↗
PubMed1992

[A study on the location of immuno-suppressive factor(s) in restraint rats and mice].

A serum lymphocyte-proliferation suppressive factor(s) induced by restraint stress over 10 h was found in previous studies in both rats and mice. The present study was undertaken to investigate the sites of its production. The results show that large doses of irradiation and cyclophosphamide (CY) decreased the total number of splenic nucleated cells, but the production of the suppressive factor was inhibited only by irradiation. This indicates that the drop in total number of lymphocytes does not play any key role in the production of the serum suppressive factor. Cell classification showed that the ratio of T to B cell was decreased by radiation but increased by CY, suggesting that this ratio may be relevant to the production of the factor. Inhibition of the production was also observed in nude mouse (an animal showing a lack of T cell activity), again supporting that T cells are involved in the production of the inhibitory factor.

Animals↗
PubMed1992

[Fatty acid compositions of cholesteryl esters. Different species and their correlation with susceptibility to atherosclerosis].

Fatty acid compositions of serum cholesteryl esters (CE) in human and animals were analysed. Polyunsaturated fatty acids (PUFA) were the predominant fatty acids in CE. A close relation between PUFA to saturated fatty acids (P/S) ratio in CE and susceptibility to atherosclerosis (AS) was observed among different species. Animals with high P/S such as tree shrews and rats (P/S 9.0) were AS-resistant, and those with low P/S such as rabbits (P/S 2.9) were susceptible to AS. The P/S ratio in human serum CE (4.9) was between those in the above two kinds of animals, and similar to those in pigs (4.2) and monkeys (3.7). A great physiological significance of abundant PUFA in CE could be expected. As the main part of low density lipoprotein (LDL), CE might provide PUFA to cells efficiently via LDL receptor. It was assumed that PUFA in CE were closely correlated with the metabolism of cholesterol and the development of AS by regulating LDL receptor activity, platelet function and prostaglandin synthesis, all of which would be affected by PUFA in cells.

Animals↗
PubMed1992

[Relationship among dosages of aminophylline, plasma levels of theophylline and variation of pulmonary arterial pressure].

The relationship among the dosages of aminophylline, plasma levels of theophylline and variations of mean pulmonary arterial pressure (mPAP) in 72 patients with COPD was investigated. The results showed that after a different loading dosage of aminophylline (6 mg/kg, 5 mg/kg and 4 mg/kg) was administered by intravenous injection, mPAP in the 6 mg/kg group was decreased more significantly (P less than 0.01) than that in the 4 mg/kg group. In the 6 mg/kg group, the decreased mPAP period sustained for 120 min, which was longer than that in the other 2 groups. The plasma levels of theophylline in the 6 mg/kg group of patients 30 to 120 min after loading dose injected were 115.54-79.04 mumol/L, which were higher than that in the others. Within the 120 min period of observation after the drug was administered no patients in any of these groups showed severe untoward effects. According to the results of this experiment, we suggest that the 6 mg/kg as a loading dose should be advised for the treatment of pulmonary hypertension in COPD. The optimum time to give the maintenance dosage should be set within 2 h after the loading dose. It is necessary to monitor the plasma levels of theophylline while aminophylline is administered, so that optimal therapeutic effects could be achieved without side effects.

Adult↗
PubMed1992

A comparative study of the outcome of renal transplantation in peritoneal dialysis and hemodialysis patients.

Fifty-four cases of renal transplantation performed in peritoneal dialysis (PD) patients were compared with 48 cases in hemodialysis (HD) patients. Three immunosuppressive treatments were used: prednisolone, azathioprine and cyclosporine. Methylprednisolone was used to treat rejection, and polyclonal Atgem or monoclonal OKT3 antibodies were used if there was no response. There was no difference in sex, age, donor source, immunosuppressive regime, duration of dialysis before transplantation, or duration of follow-up between the two groups. Following transplantation, there was no significant difference in patient mortality and survival or graft survival between the groups. The incidences of infections were also similar in the two groups. PD is commonly used in developing countries as an alternative to hemodialysis for the treatment of chronic renal failure. This study has shown that renal transplantation can be successfully performed in patients treated by this method.

Adult↗
PubMed1992

[Measurement of T-lymphocyte subsets in bronchial alveolar lavage fluid on smokers and patients with lung cancer].

The measurements of T-lymphocyte subsets (as expressed by, CD3 CD4 and CD8) in the peripheral blood and bronchial alveolar lavage fluid (BALF) on 30 cases of lung cancer, 26 cases of smoker and 25 cases of nonsmoker have been done. The results showed that there existed no differences in the peripheral blood of the above said subsets of lymphocyte. In BALF the percentage of CD3 in all the lymphocytes did not show any significant difference among the three tested groups, either (P > 0.05). But the percentage of CD4 and the ratio of CD4/CD8 manifested the following peculiarity: patients with lung cancers < smokers < nonsmokers (P < 0.01). On the contrary, the percentage of CD8 showed just the opposite: patients with lung cancer > smokers nonsmokers (P < 0.01). It was suggested that the cellular immunity as shown in the peripheral blood did not correspond with the findings in the BALF. The cellular immunity of the lungs was decreased both in patients with lung cancer and smokers, but more severely in the group of lung cancer patients. Smokers who have had changes of above said subsets of lymphocytes in the lungs may develop lung cancer. As the lung cancer advances, it may suppress body immunity and in turn enhance the cancer growth. Both the cause and the effect are interrelated. Therefore, the provocative agents for cellular immunity may act as one of the adjunctive therapies for patients with lung cancer. Giving up smoking should play an important role in recovering or promoting the immunity of the lungs and decreasing the incidence as well as improving the prognosis in patients with lung cancer.

Adult↗
PubMed1992

Structural analysis of pigeon lens crystallins by near-infrared Fourier transform Raman spectroscopy.

Crystallins from pigeon eye lenses were isolated and purified by gel-permeation and anion-exchange chromatographies and characterized by gel electrophoresis, amino acid analysis and Raman spectroscopy. alpha- and beta-Crystallins could be obtained in relatively pure forms by single-step size-exclusion chromatography whereas an extra step of ion-exchange chromatography was needed for the separation of delta crystallin from the beta-crystallin fraction. In contrast to most characterized vertebrate species, a large amount of glycogen is eluted as a high molecular form in the first peak of gel filtration column. Structural analyses of total crude soluble proteins and purified alpha-, beta- and delta-crystallin fractions were made with respect to their amino acid compositions and characteristic near-IR Fourier-transform Raman spectra. The results indicate that the major secondary structures of alpha- and beta-crystallins are mainly anti-parallel beta-pleated sheet in nature as judged by the Raman signals at 1242 (amide III) and 1669-1670 cm-1 (amide I) whereas delta-crystallin consists of a significant content of alpha-helices as evidenced by the Raman signal at 1657-1660 cm-1 (amide I). The low intensity of S-S disulfide stretching vibration at 508-510 cm-1 coupled with the presence of S-H stretching at 2560-2580 cm-1 for alpha-, beta- and delta-crystallin pointed to the fact that sulfhydryl groups in most crystallins are resistant to air oxidation during the process of homogenization and protein extraction. It is also found that the relative Raman signal intensities of Tyr, Phe, and Trp residues in purified crystallins correlate very well with the data obtained from amino acid analysis. Especially noteworthy is the demonstrated usefulness of applying Raman techniques in the detection of the microenvironments of the aromatic amino acids such as Tyr and Trp in the native crystallins, which may prove useful in the study of contribution of these aromatic residues to crystallin packing and stability.

Amino Acids↗
PubMed1992

Development and resolution of Pneumocystis carinii pneumonia in severe combined immunodeficient mice: a morphological study of host inflammatory responses.

The development and resolution of naturally-acquired Pneumocystis carinii pneumonia was studied in a severe combined immunodeficient (SCID) mouse model by light and electron microscopies. Initial infection was evident in 3-week-old SCID mice and started as focal alveolar colonization in the areas near terminal airways. Pronounced pulmonary inflammation occurred in animals of 10 weeks or older and the infection intensity reached a plateau in animals 12 weeks of age. At this stage of disease, the histopathological features of P. carinii infection in SCID mice were similar to those of immunodeficient man. Reconstitution of SCID mice with immunocompetent spleen cells at day 0 induced substantial pulmonary inflammation that was evident already by day 7 and most severe and extensive by day 12. The clearance of P. carinii did not begin until after day 12 and was almost completed by day 17. Alveolar macrophages in mice killed between days 12 and 15, at the time when P. carinii are being rapidly cleared, appeared active but phagocytosis of P. carinii was not commonly observed by either light or electron microscopy. These results suggest that (1) the presence of non-lymphoid inflammatory cells in SCID mice is not sufficient to control P. carinii infection; (2) the clearance of P. carinii from the lungs of reconstituted SCID mice requires local recruitment of large numbers of inflammatory cells with an active appearance; and (3) intracellular killing of P. carinii by phagocytosis does not appear to be a major mechanism in host defences against P. carinii infection in this model.

Age Factors↗