Geometric and electronic structure/function correlations in non-heme iron enzymes.
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Journal Article↗
Biomedical subjects
Y S Yang
Publications and source records attributed to Y S Yang.
Identification, purification, and characterization of a thermophilic imidase from pig liver.
This study investigates thermophilic imidase activity of the liver. We demonstrate that imidase catalyzes the hydrolysis of imides at a temperature substantially higher than that of its native environment. Then, a thermophilic imidase is purified to homogeneity from pig liver, and its thermoproperties are studied. About 2500-fold of purification and 15% yield of imidase activity are obtained after ammonium sulfate precipitation, octyl, DEAE, chelation, and gel filtration chromatography. While avoiding heat treatment for the protein purification, this study also indicates that only one enzyme is responsible for the imidase activity. This homogenous enzyme prefers to catalyze hydrolysis of imides at above 60 degrees C rather than at the body temperature of a pig. Although stable at below 50 degrees C, imidase quickly loses its activity at above 65 degrees C. Thus, the temperature effect on imidase activity is limited mainly by its thermostability. Substrate specificity of imidase is also temperature dependent. Our results demonstrate that the hydrolysis of physiological substrates is the most temperature dependent and that of hydantoins is the least temperature dependent. When increasing the reaction temperature from 25 to 60 degrees C, specific activities increase 50- and 60-fold for dihydrouracil and dihydrothymine, respectively. The temperature effect on the K(m) and V(max) of imidase is substrate dependent.
Amidohydrolases↗
Hormone replacement therapy reverses the decrease in natural killer cytotoxicity but does not reverse the decreases in the T-cell subpopulation or interferon-gamma production in postmenopausal women.
OBJECTIVE: To investigate the immunologic deviations of postmenopausal women before and after hormone replacement therapy (HRT). DESIGN: Prospective study. SETTING: University teaching hospital. PATIENT(S): Seventeen postmenopausal women (study group) and 17 women of reproductive age (control group). INTERVENTION(S): Continuous usage of E(2) valerate 2 mg/d and medroxyprogesterone acetate 5 mg/d in postmenopausal women in the study group. MAIN OUTCOME MEASURE(S): Immunophenotyping with flow cytometry, cytokine production with and without mitogen stimulation of the peripheral mononuclear cells, and a natural killer (NK) cell cytotoxicity test against K562 target cells by the (51)Cr-release assay were performed in the control group and in the study group before, 1 month after, and 6 months after HRT. RESULT(S): NK cytotoxicity, interferon-gamma production, and the T-cell subpopulation were significantly decreased, and the subpopulations of CD3(+)CD25(+) and CD3(+)HLA-DR(+) were increased in the study group before HRT when compared with those in the control group. After HRT was given for 6 months, however, the NK cytotoxicity increased significantly in the postmenopausal women to a value similar to that of the control group. CONCLUSION(S): Women after menopause are prone to impaired immune responses. Nevertheless, some of the impairment can be restored after HRT.
Adult↗
Cryopreservation of mature human oocytes by vitrification with ethylene glycol in straws.
OBJECTIVE: To examine the effect of vitrification with ethylene glycol (EG) for mature human oocytes in straws. DESIGN: Prospective, randomized, in vitro experiments. SETTING: Reproductive unit of a university hospital. PATIENT(S): Immature oocytes from 110 patients undergoing intracytoplasmic sperm injection (ICSI). INTERVENTION(S): The immature oocytes were incubated to reach metaphase II (MII). The MII oocytes were treated with EG-based cryoprotectants and vitrified in straws. They were diluted in sucrose solutions, inseminated by ICSI, and cultured in vitro. MAIN OUTCOME MEASURE(S): Survival, fertilization, and embryo cleavage. RESULT(S): The survival rates were greater for oocytes pretreated with 1.5 M of EG (65% for 0 minute, 93% for 5 minutes, and 96% for 10 minutes). The oocytes vitrified in 60 and 90 seconds had a greater rate of fertilization than those vitrified in 120 seconds. There were no differences in survival and fertilization for vitrified oocytes diluted by three or four steps. The cleavage rates to the six- to eight-cell stage were comparable with controls. However, no blastocyst formation was observed in vitrified oocytes. CONCLUSION(S): Vitrification of human oocytes with EG in straws achieves a high rate of survival, fertilization, and early cleavage of embryos. Further studies should be conducted for the improvement of blastocyst formation.
Cell Survival↗
Increase in the expression of killer cell inhibitory receptors on peritoneal natural killer cells in women with endometriosis.
OBJECTIVE: Malfunction of peritoneal natural killer cells (NK) may result in endometriosis. The present study was designed to determine whether the decrease in NK cytotoxicity occurs at early and advanced stages of endometriosis and is due to the increase in the NK inhibition receptors. DESIGN: A case control study. SETTING: A tertiary-care infertility center . PATIENT(S): A total of 44 women (controls, n = 11; women with early-stage endometriosis, n = 11; and women with advanced-stage endometriosis, n = 22) were included in this study. INTERVENTION(S): Laparoscopic examination. MAIN OUTCOME MEASURE(S): NK cytotoxicity was determined by assay of (51)Cr release against K562 cells, and the expression of killer cell inhibitory receptors (KIR, including NKB1, GL183, and EB6) in NK cells was examined by flow cytometry. RESULT(S): Women with endometriosis showed a decrease in peritoneal NK cytotoxicities against K562 at early and advanced stages of endometriosis. The expression of KIR (NKB1 and EB6) was significantly elevated in the peritoneal NK cells of women with advanced-stage endometriosis compared with controls. KIR (NKB1) was also significantly increased in peritoneal NK cells of women with advanced-stage endometriosis, compared with those of women with early-stage endometriosis. CONCLUSION(S): The results of this study suggest that the decrease in peritoneal NK cytotoxicities against K562 is observed and that this disease may be partially due to the increased expression of KIR on these NK cells.
Ascitic Fluid↗
Autoantigen Ro52 directly interacts with human IgG heavy chain in vivo in mammalian cells.
Previously, when we used in vivo yeast two-hybrid and in vitro protein-protein interaction analyses, we demonstrated a direct interaction between autoantigen Ro52 and the human IgG heavy chain. This interaction occurred in the absence of antibody-antigen specific interaction. Here, by employing a novel strategy, we further demonstrated that Ro52 co-localized with IgG in transfected mammalian cells. The co-localization was specific to IgG1 but not IgG3. Co-immunoprecipitating IgG with Ro52 from transfected cell lysates suggested that protein complex containing Ro52 and IgG contributed to the in vivo co-localization. In addition, IgG from normal human serum was shown to bind to the surface of apoptotic keratinocytes and the binding could be competitively blocked by 50-fold excesses of IgG1, not IgG3. With a direct binding study, we also demonstrated that IgG1 could bind to the surface of apoptotic cells while IgG3 bound barely. This binding was not competed by Fcgamma fragments indicating a non-Fcgamma receptor mediated interaction. Finally, in a competition analysis the addition of GST-RFP could reduce the IgG binding to the cell surface. Thus, we suggested that the binding of IgG to the apoptotic keratinocytes might be mediated through the interactions with the surface exposed Ro52. The potential role of forming this protein complex on the apoptotic cells will be discussed.
Animals↗
Backbone dynamics and solution structure refinement of the 15N-labeled human oncogenic protein p13MTCP1: comparison with X-ray data.
Two related oncogenes, TCL1 and MTCP1, are overexpressed in certain T-cell prolymphocytic leukemias as a result of chromosomal rearrangements that involve the translocation of one T-cell receptor gene to either chromosome 14q32 or Xq28, respectively. The human oncoprotein p13MTCP1 is coded by the MTCP1 gene and its primary sequence is highly and only homologous to that of p14TCL1, the product of TCL1. These two proteins likely represent the first members of a new family of oncogenic proteins. A previous model of the three-dimensional solution structure of p13MTCP1 was determined recently using exclusively homonuclear proton two-dimensional NMR methods and, almost simultaneously, high-resolution crystal structures of p13MTCP1 and p14TCL1 appeared in the literature. In order to gain more insight into the details of the solution structure, we uniformly labeled p13MTCP1 with nitrogen-15. The refined structure benefits from 520 additional NOEs, extracted from either 15N-edited 3D experiments or homonuclear 2D NOESY recorded at 800 MHz, and from a nearly complete set of phi angular restraints. Measurements of 15N spin relaxation times and heteronuclear 15N[1H]NOEs at two magnetic field strengths provided additional insights into the dynamics of the protein backbone. On the basis of these new results, a putative binding surface for this particular class of oncogenes is discussed.
Amino Acid Sequence↗
Nonlinear cell response to strong electric fields.
The response of living cells to externally applied electric fields is of widespread interest. In particular, the intensification of electric fields across cell membranes is believed to be responsible, through membrane rupture and reversible membrane breakdown processes, for certain types of tissue damage in electrical trauma cases which cannot be attributed to Joule heating. Large elongated cells such as skeletal muscle fibres are particularly vulnerable to such damage. Previous theoretical studies of field intensification across cell membranes in such cells have assumed the membrane current to be linear in the applied field (Ohmic membrane conductivity) and were limited to sinusoidal applied fields. In this paper, we investigate a simple model of a long cylindrical cell, corresponding to nerve or skeletal muscle cells. Employing the electroquasistatic approximation, a system of coupled first-order differential equations for the membrane electric field is derived which incorporates arbitrary time dependence in the external field and nonlinear membrane response (non-Ohmic conductivity). The behaviour of this model is investigated for a variety of applied fields in both the linear and highly nonlinear regimes. We find that peak membrane fields predicted by the nonlinear model are approximately twice as intense, for low-frequency electrical trauma conditions, as those of the linear theory.
Algorithms↗
Acarbose partially inhibits microvascular retinopathy in the Zucker Diabetic Fatty rat (ZDF/Gmi-fa).
We compared quantitative capillary retinopathic changes between non-insulin-dependent diabetic Zucker Diabetic Fatty (ZDF) rats and heterozygous nondiabetic Zucker controls and evaluated the effect of an orally administered glucosidase inhibitor, acarbose, on retinopathy in these animals. Four groups of eight rats were analyzed: treated and untreated ZDF and treated and untreated Zuckers. Retinal capillary basement membrane thickening and retinal capillary cell density were determined from transmission electron microscopy and trypsin digestion preparations. ZDF rats had thicker basement membranes (p<0.0001) and more cells per unit capillary length (p=0.0003) compared to Zuckers. Acarbose treatment significantly reduced basement membrane thickening in the treated ZDF rats (p=0.001), but the effects on cell density showed only a favorable trend. Acarbose treatment has an ameliorative effect on the development of microvascular retinopathy in the ZDF rat, probably due to lessening of hyperglycemia.
Acarbose↗
Relationships of serum pro-inflammatory cytokines and vascular endothelial growth factor with liver dysfunction in severe ovarian hyperstimulation syndrome.
The aims of this study were to determine if differences in serum pro-inflammatory cytokines, vascular endothelial growth factor (VEGF) and clinical pregnancy rate, could be observed in women with severe ovarian hyperstimulation syndrome (OHSS) in those who did and did not have liver dysfunction. Twenty-nine in-vitro fertilization patients with severe OHSS were included. The patients were divided into the normal liver function tests (LFT) group (n = 14) and the abnormal LFT group (n = 15) according to the LFT on day of admission. Periodic measurements of serum interleukin (IL)-6, IL-8, tumour necrosis factor-alpha (TNF-alpha), VEGF, oestradiol, progesterone concentrations, and LFT were performed during hospitalization. Concentrations of IL-6 in the active phase of OHSS were significantly higher in the abnormal LFT group than in the normal LFT group (19.7 +/- 15.7 versus 8.1 +/- 7.0 pg/ml, respectively). The severity of liver dysfunction was not correlated with concentrations of oestradiol, progesterone, haematocrit, white blood cell counts, or any studied cytokine. The clinical pregnancy rate was significantly lower in the abnormal LFT group (46.7%) than in the normal LFT group (85.7%). These results suggest that IL-6 cytokine system may play a role in the pathogenesis of liver dysfunction in severe OHSS. Abnormal LFT were associated with lower clinical pregnancy rates.
Adult↗
Open pulled straws for vitrification of mature mouse oocytes preserve patterns of meiotic spindles and chromosomes better than conventional straws.
Vitrification of oocytes has been applied recently for humans, but remains elusive. The microtubules of oocytes are vulnerable to cryoprotectants and thermal changes. Using mouse oocytes, the effects of vitrification in open pulled straws (OPS) were investigated on survival, the meiotic spindle, and chromosomes and compared with conventional straws. Mature oocytes were allocated to four groups for exposure to cryoprotectants, vitrification in conventional straws, or vitrification in OPS. They were diluted in stepwise sucrose solutions. Oocytes without treatments were used as controls. The surviving oocytes were stained for meiotic spindles and chromosomes. After dilution, all of the oocytes exposed to cryoprotectants survived. Vitrification sometimes resulted in lysis so that survival using OPS (62%) was significantly (P < 0.05) smaller than that using conventional straws (81%). Oocytes exposed to cryoprotectants or vitrified exhibited serious disturbances of microtubules immediately post-dilution. After 1 h incubation, the microtubules could repolymerize so that the OPS group had significantly (P < 0.05) more normal spindles (78%) than did the conventional straw group (21%). The former also tended to have more compact chromosomes (87%) than did the latter (78%). OPS for vitrification of oocytes achieve more rapid cooling, warming, and dilution and so reduce spindle injury. However, the lower survival rate in OPS needs improvement.
Animals↗
Value of serum and follicular fluid cytokine profile in the prediction of moderate to severe ovarian hyperstimulation syndrome.
The aim of this study was to examine the role of serum and follicular fluid pro-inflammatory cytokines and vascular endothelial growth factor (VEGF) in the prediction of ovarian hyperstimulation syndrome (OHSS). A total of 156 consecutive women undergoing in-vitro fertilization were recruited. The study group comprised 12 women who subsequently developed moderate (n = 7) or severe (n = 5) OHSS. The two control groups were comprised of a randomized selection of 12 high-risk and 12 low-risk women in whom OHSS did not develop. Serum was collected on days of human chorionic gonadotrophin, oocyte retrieval, and embryo transfer. Serum and follicular fluid concentrations of interleukin (IL)-6, IL-8, tumour necrosis factor-alpha (TNF-alpha), and VEGF were measured. Follicular fluid IL-6 concentrations at the time of oocyte retrieval and serum IL-8 concentrations at the time of embryo transfer were significantly higher in the OHSS compared to the two control groups (P = 0.026 and P = 0.017 respectively). Serum concentrations of TNF-alpha and VEGF showed no statistically significant difference between the OHSS group and the controls at any studied time point. This study suggests that follicular fluid IL-6 concentrations at the time of oocyte retrieval and serum IL-8 concentrations on the day of embryo transfer may serve as early predictors for this syndrome.
Adult↗
Fuzzy EMG classification for prosthesis control.
This paper proposes a fuzzy approach to classify single-site electromyograph (EMG) signals for multifunctional prosthesis control. While the classification problem is the focus of this paper, the ultimate goal is to improve myoelectric system control performance, and classification is an essential step in the control. Time segmented features are fed to a fuzzy system for training and classification. In order to obtain acceptable training speed and realistic fuzzy system structure, these features are clustered without supervision using the Basic Isodata algorithm at the beginning of the training phase, and the clustering results are used in initializing the fuzzy system parameters. Afterwards, fuzzy rules in the system are trained with the back-propagation algorithm. The fuzzy approach was compared with an artificial neural network (ANN) method on four subjects, and very similar classification results were obtained. It is superior to the latter in at least three points: slightly higher recognition rate; insensitivity to overtraining; and consistent outputs demonstrating higher reliability. Some potential advantages of the fuzzy approach over the ANN approach are also discussed.
Algorithms↗
Luteinizing hormone up-regulates the expression of interleukin-1 beta mRNA in human granulosa-luteal cells.
PROBLEM: Previously, we observed that follicular fluid obtained from patients with premature luteinization contained elevated interleukin-1 beta (IL-1 beta) levels. In this study. we aimed to examine the effects of luteinizing hormone (LH) on IL-1 beta expression and IL-1 beta on steroidogenesis in human granulosa-luteal cells. METHOD OF STUDY: Human granulosa-luteal cells were obtained during oocyte retrieval. The cells were treated with either LH or IL-1 beta and subsequently were examined for the level of IL-1 beta transcript. The conditioned media were examined for IL-1 beta protein and steroid hormone levels. RESULTS: LH (250-500 mIU/mL) up-regulated the expression of IL-1 beta mRNA (up to a 4-fold increase over control; P<0.05) in the granulosa-luteal cells. IL-1 beta (5-50 ng/mL) increased the basal, but not LH-dependent, progesterone production from these cells in a dose-dependent manner after 96 and 144 hr of culture (P<0.05). However, an inhibitory effect of IL-1 beta on LH-dependent estradiol production was observed (up to 20% decrease, P<0.05). CONCLUSIONS: LH is capable of stimulating IL-1 beta transcript expression in human granulosa-luteal cells and may regulate ovarian steroidogenesis, at least partly through the activation of IL-1 beta.
Cells, Cultured↗
NRAMP1 gene polymorphisms in patients with rheumatoid arthritis in Koreans.
Natural resistance-associated macrophage protein 1 (Nramp1) is a genetic locus associated with innate resistance or susceptibility of murine hosts to infection with intracellular pathogens such as Salmonella, Leishmania and Mycobacterium. The human homologue of the Nramp1 gene, designated NRAMP1, has been investigated as a candidate gene for genetic susceptibility to autoimmune diseases as well as infections. This study tries to determine whether NRAMP1 polymorphisms are associated with susceptibility to rheumatoid arthritis in Koreans. The nine NRAMP1 polymorphisms (1 microsatellite, 1 variation in 3' UTR, 5 silent substitution, 2 amino acid substitution) were typed by PCR-RFLP in 74 patients with rheumatoid arthritis (RA) and 53 healthy controls in Koreans. The distribution of allele and genotype frequencies were compared between patients and controls. Three NRAMP1 polymorphisms (823C/T, D543N and 1729+55del4) were significantly associated with RA. In addition, there were significant differences in the genotype frequencies for 823C/T, D543N and 1729+ 55del4 polymorphisms between RA patients and controls. Genotypes of A/A homozygote for D543N and TGTG deletion homozygote for 1729+55del4 were only detected in the patient group. These data indicate that genetic polymorphisms of NRAMP1 might be associated with the susceptibility to rheumatoid arthritis in Koreans.
Alleles↗
Enterobius vermicularis egg positive rate of pre-school children in Chunchon, Korea (1999).
Pre-school children were examined for the presence of Enterobius vermicularis egg by perianal swab method. We visited 67 nurseries and 25 kindergartens between January, 1999 and April, 1999 in Chunchon, Korea. Of the 4,711 children examined, 434 (9.2%) were found to be positive for E. vermicularis egg. The egg positive rates of boys and girls were 10.1% and 8.1%, respectively. The rates were 7.8% in kindergartens and 9.7% in nurseries. Positive rate of 50 institutions was less than 10%. Rate in 35 institutions was from 10% to less than 20%. Rate in 7 institutions was equal to or over 20%. Out of 1,113 children examined twice, 28 (2.5%) children were positive consecutively, 53 (4.8%) were positive with negative conversion, 47 (4.2%) were negative with positive conversion, and 985 (88.5%) were consecutively negative. It means that in the low endemic area of enterobiasis with around 10% positive rate, the two consecutive examinations may increase the egg detection rate of 4.2-4.8%. The small number of consecutive egg positive children also suggests that the worm burden of the positive children might be low. Since the egg positive rate of that age group in this City increased from 1.85% (1997), and 3.0% (1998), the more intensive regular control should be executed.
Animals↗
Diagnosis and follow-up of acute promyelocytic leukemia by detection of PML-RAR alpha gene rearrangement.
BACKGROUND: Rapid and accurate diagnosis of acute promyelocytic leukemia (APL) is essential for management of the disease, as all-trans retinoic acid (ATRA) therapy only induces complete remission in patients whose leukemic cells harbor a t(15;17) translocation, resulting in promyelocytic-retinoic acid receptor alpha (PML-RAR alpha) fusion transcripts. Moreover, a positive reverse transcriptase-polymerase chain reaction (RT-PCR) of PML-RAR alpha is reported to be a sensitive predictor of relapse in APL. This prompted us to use RT-PCR for rapid diagnosis and monitoring of minimal residual disease in APL patients. METHODS: A nested RT-PCR technique was applied to detect the unique PML-RAR alpha fusion transcript in 13 APL patients. The test was applied to help clarify the diagnosis and monitor minimal residual disease after treatment. RESULTS: All 13 APL patients had a positive test result: five patients with the S-form, seven patients with the L-form and one patient with the V-form of mRNA fusion transcripts. Minimal residual disease was prospectively monitored using this technique in six patients. Although in clinical remission, all four patients treated with ATRA alone were persistently PCR positive. Of the six patients receiving various forms of consolidation chemotherapy, one was persistently PCR positive while in remission and relapsed four months after the positive PCR test. Five patients were PCR negative. One of the five negative patients relapsed six months after a negative PCR test. The other four patients remained in remission, with a follow-up period of 25 to 46 months after the negative test. PCR was performed in two patients who had been in continuous remission for 3.5 and seven years, respectively. They both had negative PCR tests. CONCLUSIONS: Nested RT-PCR is valuable for confirming the diagnosis of APL and in monitoring minimal residual disease. However, we found that negative test cannot absolutely exclude the possibility of future relapse.
Adolescent↗
A cooperative model for Ca(++) efflux windowing from cell membranes exposed to electromagnetic radiation.
We propose a simplified version of a cooperative lattice membrane model given by Grodsky to explain observed Ca(++) efflux windowing effects from cell membranes exposed to electromagnetic radiation. Assuming that field induced conformational interactions occur only between bistable receptor sites and glycoprotein Ca(++) sites on the surface of the membrane, the model is shown to be equivalent to an Ising model. This model is known to have a phase transition to an ordered state in which a macroscopic number of Ca(++) sites are either occupied or unoccupied. We identify such states with enhanced Ca(++) efflux from cell membranes. By further assuming an averaged signal, sinusoidally varying coupling between receptor and Ca(++) sites and a power-law dependence of the characteristic time constant on the induced power-density of the applied field, we show that the model is consistent with published experimental results on power density windowing effects for particular values of model parameters. For these parameter values, the model predicts further power densities where windowing effects may be observed under appropriate conditions.
Algorithms↗