Search PubMed⌕ Search

Biomedical subjects

D Y Chen

Publications and source records attributed to D Y Chen.

At least 37 records · Page 2Linked to original sources
PubMed2001

Refrigeration of donor cells in preparation for bovine somatic nuclear transfer.

In mammals, preparation of donor cells for somatic nuclear transfer is very important because the character of the donor cell directly affects the efficiency and outcome of transfer. The protocols used most commonly for donor preparation are (i) disaggregating cells from fresh tissue 1-2 h before micromanipulation or (ii) trypsinizing cultured cells temporarily, after special treatments for 3-8 days (for example, serum starvation). In this study, a new simple protocol was designed, whereby the donor cells (cumulus cells) used in bovine somatic nuclear transfer were refrigerated. In brief, cultured cells at 80-100% confluency were detached using trypsin, washed by centrifugation, aliquoted into different vials and refrigerated at 4 degrees C. The density of viable cells was decreased after day 1 of refrigeration; however, the rate of decrease tended to slow down with increasing duration of refrigeration. Cells refrigerated for 15 days were seeded at a density of 5 x 10(4) ml(-1) and reached 70% confluency after day 2 of culture. Most cells had the normal number of chromosomes (2n = 60). Cells chilled at 4 degrees C for different durations were removed from refrigeration and immediately subjected to micromanipulation. The in vitro development of reconstructed embryos (fusion rates, cleavage rates, morula and blastocyst rates) indicated that there were no significant differences among treatment groups regardless of the duration of refrigeration (0-2 weeks) of the donor cells. Reconstructed embryos were transferred into the uteri of recipient cows. No significant differences were observed in established early pregnancies between embryos derived from the non-refrigerated donor cells and those derived from refrigerated donor cells. This study indicates that refrigeration of donor cells for 1-2 weeks is a feasible protocol for preparing donor cells for bovine somatic nuclear transfer, and does not compromise development in vitro and early development in vivo.

Animals↗
PubMed2000

Effect of telophase enucleation on bovine somatic nuclear transfer.

Telophase enucleation has been proven to be an efficient method for preparing recipient cytoplasts in bovine embryonic nuclear transfer (2, 11). This research was designed to study in vitro development of bovine oocytes containing transferred somatic cell nuclei, reconstructed by using enucleated in vitro-matured oocytes 32 h of age at telophase II stage as recipient cytoplasts, compared with those 24 h of age at metaphase II stage. Two protocols for donor cell injection were adopted, i.e., subzonal injection (SUZI) and intracytoplasmic injection (ICI). Bovine oviduct epithelial cells (BOECs) and bovine cumulus cells (BCCs) from an adult cow were used as nuclear donors for these experiments. In SUZI groups, the fusion rate of donor cells, both BOECs and BCCs, with MII enucleated oocytes were higher than those with TII enucleated oocytes (54% vs. 41% and 53% vs. 39%, respectively; P<0.05), but the development rates to morula plus blastocyst stage in MII groups were lower than those in TII groups (22% vs. 39% and 21% vs. 41%, respectively; P<0.05). In ICI groups, about 26% of enucleated MII oocytes injected with BOECs or BCCs cleaved and only small parts of them developed to blastocyst stage (4% and 3%, respectively; P>0.05). When BOECs or BCCs were intracytoplasmically injected into oocytes enucleated at TII stage, no blastocyst was formed in either donor cell group and no cleavage occurred in BOEC group. Our data demonstrated that telophase enucleation is beneficial to early embryo development when bovine somatic nuclei are transferred by subzonal injection. However, it is harmful when donor cells are directly injected into the cytoplast of the enucleated oocytes.

Animals↗
PubMed2000

Studies in marine polypropionate synthesis: total synthesis of (-)-baconipyrone C.

[reaction--see text] An asymmetric total synthesis of the unusual siphonariid metabolite, (-)-baconipyrone C (3), is described. Key steps included a tin(II)-mediated aldol coupling for the preparation of the carboxylic acid 17 and two different boron-mediated aldol additions leading to alcohol 8. Ester formation using modified Yamaguchi conditions gave 24, leading on PMB deprotection to (-)-baconipyrone C.

Animals↗
PubMed2000

Startle responses to electric shocks: measurement of shock sensitivity and effects of morphine, buspirone and brain lesions.

The present study developed a new protocol to assess shock sensitivity in rats. Male Wistar rats were subjected to footshock stimuli ranging from 0 to 1.6 mA (0.1 s) in a startle apparatus and startle responses elicited by shocks were measured. Acoustic stimuli (95, 105, or 115 dB) were dispersed within the shock series serving as a control measurement of motor performance. Results indicated that the magnitude of shock startle responses significantly increased with the shock intensity in a linear trend. Morphine (8.0 mg/kg) and buspirone (1.0, 2.5, or 5.0 mg/kg), both of which possessing analgesic effects, depressed shock startle but had no such effect on acoustic startle. The effect of morphine was readily reversed by pretreatment of naloxone (1.0 mg/kg). To investigate the neural basis underlying this response, radio-frequency lesions of various structures implicated in processing of nociceptive or aversive information were undertaken. Lesions of the ventroposterior thalamic nucleus, insular cortex, or amygdala decreased startle reactivity to electric shocks but not to acoustic stimuli. Lesions of the anterior cingulate gyrus or medial prefrontal cortex, while altered the reactivity to acoustic stimuli, had no effect on the shock-elicited startle. These results suggested that the amplitude of startle in response to electric shocks provide a quantitative measurement of shock sensitivity within an extended range of stimulus intensities. Performing this response may engage the the central nociceptive pathway.

Animals↗
PubMed2000

Trichosanthin induced calcium-dependent generation of reactive oxygen species in human choriocarcinoma cells.

The type-I ribosome-inactivating protein trichosanthin (TCS) has a broad spectrum of biological and pharmacological activities, including abortifacient, anti-tumor and anti-HIV. We found for the first time that TCS induced the generation of reactive oxygen species (ROS) in human choriocarcinoma cells (JAR cells) at the level of the single cell by using the fluorescent probe 2',7'-dichlorofluorescein diacetate with confocal laser scanning microscopy. TCS-induced ROS formation was shown to be dependent on the presence of extracellular Ca2+ and was further reduced when cytosolic Ca2+ was chelated by BAPTA-AM. The production of ROS increased rapidly after the application of TCS, which paralleled TCS-induced increase in intracellular calcium monitored using fluo 3-AM. Simultaneous observation of the nuclear morphological changes via two-photon laser scanning microscopy and production of ROS via confocal laser scanning microscopy revealed that ROS were involved in the apoptosis of JAR cells. The contribution of ROS was confirmed by experiments in which the antioxidant alpha-tocopherol prevented TCS-induced ROS formation and cell death. The finding that TCS induced calcium-dependent generation of ROS in JAR cells and that ROS were involved in the apoptosis of JAR cells might provide new insight into the anti-tumor and anti-HIV mechanism of TCS.

Antineoplastic Agents, Phytogenic↗
PubMed2000

ExoT of cytotoxic Pseudomonas aeruginosa prevents uptake by corneal epithelial cells.

The presence of invasion-inhibitory activity that is regulated by the transcriptional activator ExsA of cytotoxic Pseudomonas aeruginosa has previously been proposed. The results of this study show that both ExoT and ExoS, known type III secreted effector proteins of P. aeruginosa that are regulated by ExsA, possess this activity. Invasion was reduced 94.4% by ExoT and 96.0% by ExoS. Invasion-inhibitory activity is not linked to ADP-ribosylation activity, at least for ExoS, since a noncatalytic mutant also inhibits uptake by an epithelial cell line (invasion was reduced 96. 0% by ExoSE381A).

Animals↗
PubMed2000

Economic and outcomes assessment of magnetic resonance imaging in the evaluation of headache.

We sought to evaluate the economic impact and diagnostic utility of magnetic resonance imaging (MRI) in the management of patients with headache and nonfocal physical examinations. Computerized medical records were retrospectively reviewed of 1,233 patients presenting for MRI of headache at our institution over a 3-year period (1992-1995). Patients with focal findings at physical examination, prior brain surgery, head trauma, or immunocompromise were excluded. A model was developed to assess the cost associated with the MR test results, and actual average institutional costs of performing an examination applied. Correlative statistical analysis of referring specialties and positive tests was also performed. Three hundred twenty-eight patients who met the above criteria were retained in the sample. One hundred sixty-three patients (50%) had negative MR test results. Of the 50% of patients with positive studies, only 5 (1.5%) had clinically significant MR results. The average cost of an MR examination was 517 dollars (1998 dollars). The cost per clinically significant managed case detected was 34,535 dollars. No statistically significant difference was found among referring specialties and clinically significant MR results. Our results indicate that MRI of nonfocal headache yields a low percentage of positive clinically significant results and has limited cost-effectiveness. Referring specialty had no significant bearing on these outcomes, regardless of specialist experience.

Adolescent↗
PubMed2000

[Determination of benzene, toluene and xylene in marine sediment samples by micellar electrokinetic capillary chromatography].

The analysis and determination of the contents of benzene series compounds in marine sediment are important in chemical exploration of petroleum resources. In the present study a method has been developed for this purpose by using micellar electrokinetic capillary chromatography (MECC). A fused silica capillary of 45 cm x 75 microns i.d. (effective length 40 cm) was used. Running buffer was 2 mmol/L aqueous solution of sodium tetraborate containing 75 mmol/L sodium dodecyl sulfate (pH 9.16) and 20% methanol. The applied voltage was 25 kV and detection wavelength was 200 nm. The benzene, toluene and xylene were separated well within 20 min. Peak area was used for quantitative analysis. The calibration curves were linear in the range 4-50 mg/L. The relative standard deviation was within 6.2%. The contents of benzene series in sediments were between 3.79 and 17.36 micrograms/g.

English Abstract↗
PubMed1999

CAMP inhibits mitogen-activated protein (MAP) kinase activation and resumption of meiosis, but exerts no effects after spontaneous germinal vesicle breakdown (GVBD) in mouse oocytes.

Various signaling molecules have been implicated in the oocyte G2/MII transition, including protein kinase C (PKC), cAMP and mitogen-activated protein (MAP) kinases. However, the cross-talk among these signaling pathways has not been elucidated. The present study demonstrates that both germinal vesicle break down (GVBD) and MAP kinase phosphorylation (activation) are inhibited when intraoocyte cAMP is increased by treating the GV-intact oocytes with dibutyryl cyclic AMP (dbcAMP), forskolin, or isobutylmethylxanthine (IBMX). Okadaic acid, a specific inhibitor of protein phosphatase-1 and -2A, completely overcame this effect. Calphostin C, a specific inhibitor of PKC, accelerated both GVBD and MAP kinase phosphorylation, and this effect was attenuated by increased intraoocyte cAMP, whereas PKC activation inhibited these events. Once GVBD occurred, the progression of oocyte maturation and MAP kinase phosphorylation were independent of cAMP These results indicate that an increase in intraoocyte cAMP, in synergy with PKC activation, initiates a cascade of events resulting in inhibition of MAP kinase phosphorylation and GVBD in the mouse oocyte.

1-Methyl-3-isobutylxanthine↗
PubMed1999

Subcallosal striations: early findings of multiple sclerosis on sagittal, thin-section, fast FLAIR MR images.

PURPOSE: To determine how sensitive and specific the magnetic resonance (MR) imaging finding of subcallosal striations is for multiple sclerosis (MS). MATERIALS AND METHODS: In 18 patients with clinically suspected MS and 32 age-matched patients without MS, sagittal 2-mm fast fluid-attenuated inversion-recovery (FLAIR) imaging was added to the routine MR studies of the brain. The images were reviewed for the presence of subcallosal striations, that is, linear, 1-mm-thick foci of hyperintensity perpendicular to the ependyma, like a stack of coins. The images were masked to exclude the hemispheric white matter and were interpreted without knowledge of patient age or medical history. RESULTS: Seventeen patients with clinical MS had subcallosal striations; one did not. Of the 32 patients without MS, five had subcallosal striations and 27 did not. Subcallosal striations were highly associated (P < .001) with clinical MS. CONCLUSION: Subcallosal striations are thought to represent perivenular demyelination, that is, the same process that later produces the "ovoid" lesions visible on routine MR images. Subcallosal striations are not seen on routine axial MR images and can be seen only on 2-mm sagittal FLAIR images.

Adult↗
PubMed1998

Functional magnetic resonance imaging in macaque cortex.

The ability to use fMRI in a monkey model would bridge the gap between the fMRI demonstration of cerebral activation in humans and the cumulative wealth of monkey data on the functional organization of the brain from single electrode mapping, radioisotope and histology studies. We report a new technique for fMRI in an awake co-operative rhesus macaque (Macaca mulatta) in a conventional clinical 1.5T MR scanner and present the first fMRI images from a macaque. Good resolution, signal-to-noise ratio and BOLD response (2.6-4.6%) have been achieved using the manufacturer's standard volume knee coil. T1 values of macaque gray and white matter (1490 ms, 1010 ms respectively) are higher than human brain, whereas T2 values are lower (55 ms, 48 ms respectively). An MR-compatible design for restraining the monkey is also described, along with a suitable EPI sequence for BOLD images, optimized for monkey T2, with voxel sizes from 29 to 61 microl, and MPRAGE sequence for anatomical studies with 0.8 mm isotropic resolution, optimized for monkey T1.

Animals↗
PubMed1998

Osteoporosis in Colles fracture.

Previous studies of bone density in patients with Colles fracture have been inconclusive. We measured bone density at the second metacarpal in 36 postmenopausal women with Colles fracture. The patients were found to have highly significant (P < 0.01) or significant (P < 0.05) differences in the parameters of bone density compared with controls matched for age. The results suggest that osteoporosis is of definite relevance to the etiology of Colles fracture in postmenopausal women.

Aged↗
PubMed1998

Experience with laser hair transplantation in Orientals.

BACKGROUND: The advent of high-energy carbon dioxide (CO2) laser-assisted hair transplantation in recent years provides a new recipient area technique for hair restoration surgery. It offers the advantages of less bleeding, time consumption, and graft compression. Specific laser parameters have been recommended in Caucasian but not Oriental patients. OBJECTIVE: To determine the effectiveness as well as appropriate parameters of laser hair transplantation in Oriental patients. METHODS: A Sharplan SilkTouch laser with the Flashscanner set to produce a 0.8-1-mm recipient site hole was employed for all subjects. Various CO2 laser parameters were used to compare the graft survival rate with traditional Nokor needle or microdilator methods for micrografting on each side of the scalp. A total of 20 patients joined the study in the past 18 months. A skin biopsy was performed to examine the tissue effect of the different laser parameters. RESULTS: Hair survival was less than that seen with conventional micrografting techniques in the first five cases, in whom CO2 laser parameters of 36-40 W and 0.2-0.7 seconds were used. After adjusting the parameters to 55 W and 0.1-0.15 seconds, the survival rate increased to between 80% and 90%. In addition, most patients thought the laser-prepared sites resulted in a more natural appearance and better hair density, when they were compared with conventionally prepared sites. CONCLUSIONS: From our study, we find that CO2 laser parameters of 55 W and 0.1-0.15 seconds seem to be the appropriate parameters for Oriental hair transplant patients. Further studies are required to establish laser parameters that will produce better hair survival.

Alopecia↗
PubMed1998

Development of recombinant human prolactin receptor antagonists by molecular mimicry of the phosphorylated hormone.

Previous studies have demonstrated that naturally phosphorylated PRL antagonizes the growth-promoting effects of unmodified PRL in two different PRL-responsive cell lines. In this study our aim was to produce a molecular mimic of phosphorylated PRL by substituting a fairly bulky, negatively charged amino acid (glutamate or aspartate) for the normally phosphorylated serine [serine 179 in human PRL (hPRL)]. In addition, because of the marked effect of phosphorylation on biological activity, we investigated the importance of the unmodified serine in the growth-promoting activity of PRL. hPRL complementary DNA was obtained from the American Type Culture Collection and subcloned into pT7-SCII after site-directed mutagenesis using the deoxyuridine approach. Proteins were expressed in Escherichia coli BL21 (DE3) and were primarily found in inclusion bodies. Agonist and antagonist activities of each serine 179 mutant were assessed using the Nb2 bioassay. Compared with standard hPRL, the recombinant wild-type was more active in the Nb2 assay, attesting to both the absence, or low level, of endotoxin contamination in preparations from these cells and the appropriate folding of the molecule. The aspartate and glutamate mutants had no intrinsic agonist activity, but both antagonized the growth-promoting activity of wild-type PRL, with the aspartate mutant proving to be a very effective antagonist. Two hundred picograms per ml of the aspartate mutant negated 75% of the growth response to 400 pg/ml wild-type PRL. When serine 179 was mutated to alanine or valine, mutant PRLs with 0% and 14% of the biological activity of wild-type PRL, respectively, were produced. These results demonstrate 1) that molecular mimicry of the phosphorylated hormone does produce a PRL antagonist, and 2) that the serine at position 179 is crucial to the growth-promoting activity of PRL. The aspartate mutant can now be used to study many aspects of the physiology of PRL.

Amino Acid Sequence↗
PubMed1998

Hibernoma. Case report and literature review.

BACKGROUND: Hibernoma is a rare type of benign tumor that originates in brown fat. It has been rarely described in dermatologic literature. OBJECTIVE: To report and discuss the clinical and histologic features of a patient with hibernoma and review the relevant literature. PATIENT: A large palpable hibernoma located in the interscapular area of a 41-year-old female was identified by physical examination. The tumor was a brown soft mass measuring 7.0 x 6.0 x 3.0 cm that involved subcutaneous tissue and was embedded in the trapzius muscle. It was a well encapsulated and highly vascular tumor. Histologic examination revealed characteristic multivacuolated oval and polygonal cells with eosinophilic, granular cytoplasm. CONCLUSION: A hibernoma can be diagnosed on the basis of gross physical characteristics and microscopic findings. This is a benign tumor with no malignant potential. Excisional biopsy is indicated to differentiate hibernoma from well-differentiated liposarcoma. Complete excision is the treatment of choice.

Adult↗
PubMed1997

The von Hippel-Lindau tumor-suppressor gene product forms a stable complex with human CUL-2, a member of the Cdc53 family of proteins.

The inactivation of the von Hippel-Lindau (VHL) gene predisposes affected individuals to VHL syndrome and is an early genetic event associated with sporadic renal cell carcinoma and CNS hemangioblastomas. The VHL protein (pVHL) has been shown to form a stable complex with elongin B and elongin C, two factors that stabilize and activate the transcription elongation factor elongin A. Here, Hs-CUL-2, a member of the recently identified multigene family, the cullins, is shown to specifically associate with the trimeric pVHL-elongin B-C (VBC) complex in vitro and in vivo. Nearly 70% of naturally occurring cancer-predisposing mutations of VHL disrupt this interaction. The pVHL-Hs-CUL-2 association is strictly dependent on the integrity of the trimeric VBC complex. Immunofluorescence studies show Hs-CUL-2 to be a cytosolic protein that can be translocated to the nucleus by pVHL. Recently it has been shown that a yeast Hs-CUL-2 homolog, Cdc53, is part of a ubiquitin protein ligase complex that targets cell cycle proteins for degradation by the ubiquitin proteolytic pathway. In Caenorhabditis elegans, a null mutation of another Hs-cul-2 homolog, Ce-cul-1, results in hyperplasia in all tissues and is required for cell cycle exit. Hence, Hs-cul-2 may be required for VHL function and, therefore, may be a candidate human tumor-suppressor gene.

Amino Acid Sequence↗
PubMed1997

Activation of protein kinase C induces cortical granule exocytosis in a Ca(2+)-independent manner, but not the resumption of cell cycle in porcine eggs.

The effects of protein kinase C (PKC) activation on meiotic resumption and cortical granule (CG) exocytosis as well as its dependence on Ca2+ in porcine eggs matured in vitro were studied. Cortical granule release was judged by both confocal laser microscopy after the eggs were labeled with fluorescein isothiocyanate-peanut agglutinin (FITC-PNA) and electron microscopy. Meiotic resumption and pronuclear formation were observed after eggs were stained with acetic orcein. When eggs were treated with PKC activators, 1-oleyl-2-acetyl-glycerol (OAG) or phorbol 12-myristate 13-acetate (PMA), the pronuclear formation percentage was significantly lower than that of Ca2+ ionophore A23187-treated group, but not statistically different from that in negative control group (P > 0.05), and most of the eggs were still arrested at metaphase II stage, suggesting that PKC activation does not induce the resumption of meiosis and pronuclear formation. In contrast, PKC activation induced 89.1% to 100% of the eggs completely or partially released their CG in different groups, not statistically different from A23187-treated group, and this effect could be overcome by PKC inhibition. When the intracellular free Ca2+ was chelated with acetoxymethal ester form of 1,2-bis(O-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid (BAPTA-AM), and then treated with PMA or OAG in Ca(2+)-free medium, the proportions of eggs with CG release were 90.9% and 78.1%, respectively, not statistically different from the above-treated groups, suggesting that CG exocytosis induced by PKC activation is independent of Ca2+ rise. The results indicate that different events of porcine egg activation may be uncoupled from one another.

Animals↗