[International expert consensus on gene therapy for hereditary hearing loss: based on clinical trials].
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Journal Article↗
Biomedical subjects
H Wu
Publications and source records attributed to H Wu.
Molecular genetic analysis of bacteriophage P22 gene 3 product, a protein involved in the initiation of headful DNA packaging.
Bacteriophage P22 DNA packaging events occur in processive series on concatemeric phage DNA molecules. At the point where such series initiate, the DNA is recognized at a site called pac, and most molecular left ends are generated within six short regions called end sites, which are present in a 120 base-pair region surrounding the pac site. The bacteriophage P22 genes 2 and 3 proteins are required for successful generation of these ends and DNA packaging during progeny virion assembly. Mutants lacking the 162-amino-acid gene 3 protein replicate DNA and assemble functional procapsids. In this report we describe the nucleotide changes and DNA packaging phenotypes of a number of missense mutations of gene 3, which give the phage a higher than normal frequency of generalized transduction. In cells infected by these mutants, more packaging events initiate on the host chromosome than in wild-type infections, so the mutations are thought to affect the specificity of packaging initiation. In addition to having this phenotype, these mutations affect the process of phage DNA packaging in detectable ways. They may: (1) alter the target site specificity for packaging; (2) make target site recognition more promiscuous; (3) affect end site utilization; (4) alter the pac site; and (5) cause apparent random DNA packaging series initiation on phage DNA.
Bacteriophage P22↗
Regulated expression of the human acetylated low density lipoprotein receptor gene and isolation of promoter sequences.
The acetylated low density lipoprotein (AcLDL) receptor is expressed on tissue macrophages after their differentiation from monocyte precursors and has been proposed to play a role in the generation of foam cells in atherosclerotic lesions. In the present studies, THP-1 human monocytic leukemia cells were used to investigate mechanisms responsible for expression of the AcLDL receptor gene after treatment with phorbol 12-myristate 13-acetate (TPA). TPA-dependent accumulation of AcLDL receptor mRNA was not detected until after a lag phase of 12 hr and was blocked by concurrent treatment with cycloheximide. In addition, the TPA-dependent induction of AcLDL receptor activity and mRNA levels was inhibited by retinoic acid and dexamethasone treatment. Isolation and sequence analysis of the promoter regions for the human and bovine AcLDL receptor genes indicated high sequence similarity. Binding sites for AP-1 proteins or other known transcription factors were not conserved between the two species, suggesting that novel factors are required for AcLDL receptor expression.
Base Sequence↗
Direct relationship between intracellular calcium mobilization and phospholipase D activation in prostaglandin E-stimulated human erythroleukemia cells.
The relationship between calcium mobilization and phospholipase D (PLD) activation in response to E-series prostaglandins (PGEs) was investigated in human erythroleukemia cells. Intracellular free Ca2+ concentration ([Ca2+]i) was increased by PGE1 and PGE2 over the same concentration range at which PLD activation was seen. Pretreatment of cells with pertussis toxin greatly inhibited the PGE-stimulated increase in [Ca2+]i, implying that a G protein participates in the PGE receptor signaling process. The peak level and also the plateau level of Ca2+ mobilization stimulated by these prostaglandins were markedly decreased in Ca(2+)-depleted medium, indicating that both extracellular and intracellular Ca2+ stores contribute to the changes in [Ca2+]i. Likewise, activation of PLD by PGE1 and PGE2 was abolished by pertussis toxin pretreatment or incubation in Ca(2+)-depleted medium. U73122, a putative phospholipase C inhibitor, blocked both Ca2+ mobilization and PLD activation in PGE-stimulated cells. Furthermore, the intracellular loading of BAPTA, a Ca2+ chelator, inhibited both Ca2+ mobilization and PLD activation by PGE1 and PGE2 in a similar dose-dependent manner. Simultaneous measurement of [Ca2+]i and PLD activity in the same cell samples indicated that PLD activity increases as a function of [Ca2+]i in a similar fashion in cells stimulated either by PGEs or by the calcium ionophore ionomycin. Taken together, these findings suggest that a rise in [Ca2+]i is necessary for PGE-stimulated PLD activity in human erythroleukemia cells.
Alprostadil↗
Structure determination of monoclinic canine parvovirus.
The three-dimensional structure of the single-stranded DNA canine parvovirus has been determined to 3.25 A resolution. Monoclinic crystals belonging to space group P2(1) (a = 263.1, b = 348.9, c = 267.2 A, beta = 90.82 degrees) were selected for data collection using primarily the Cornell High Energy Synchrotron Source and oscillation photography. There was one icosahedral particle per crystallographic asymmetric unit, giving 60-fold redundancy. The particle orientations in the unit cell were determined with a rotation function. The rough positions of the particles in the unit cell were estimated by considering the packing of spheres into the P2(1) crystal cell. More accurate particle centers were determined from Harker peaks in a Patterson function. Hollow-shell models were used to compute phases to 20 A resolution. The radii of the models were based on packing considerations, the fit of spherical shells to the low-resolution X-ray data and low-angle solution scattering data. The phases were extended to 9 A resolution using molecular replacement real-space averaging. These were then used to determine the heavy-atom position of a K2PtBr6 derivative, for which only 5% of the theoretically observable reflections had been recorded. The center of gravity of the 60 independent heavy-atom sites gave an improved particle center position. Single isomorphous replacement phases to 8 A resolution were then calculated with the platinum derivative. These were used to initiate phase improvement and extension to 3.25 A resolution using density averaging and Fourier back-transformation in steps of one reciprocal lattice point at a time. The resulting electron density map was readily interpretable and an atomic model was built into the electron density map on a PS390 graphics system using the FRODO program. The R factor prior to structure refinement for data between 5.0 and 3.25 A was 36%.
Crystallization↗
Analysis of the biocompatibility of magnetic porous tricalcium phosphate ceramics in rat femurs.
A new biomaterial, magnetic porous tricalcium phosphate ceramics (MPTCP), may be used for repairing defects in bones as a substitute for bone grafting. This study was designed to examine its biocompatibility and the systemic toxicity. The ceramic sections, approximately 15.3 mg, were implanted into both proximal femurs of 10 male and 10 female albino SW rats. An equal number of male and female animals served as sham-operated controls. Implantation of MPTCP ceramics in rat femurs had no deleterious effects on body weight, organ/body weight ratio, and on blood, liver and kidney functions. Plasma levels of inorganic phosphorus were significantly elevated in rats implanted with MPTCP ceramics. Alkaline phosphatase values in plasma in the experimental group were equal to those in the control group. Photomicrographs, radiographs and scanning electron photomicrographs of the rat femurs implanted with MPTCP ceramics demonstrated excellent bone association with implants and growth of some new bone into MPTCP ceramics. In conclusion, MPTCP ceramics are biocompatible and suitable for the reconstruction of bones.
Animals↗
Porcine endometrial epithelial cells immortalized by transfection with origin-defective, temperature-sensitive simian virus 40 DNA.
The purpose of this study was to immortalize porcine endometrial cells and to characterize the transformed cells. Primary porcine endometrial cells were transfected with the plasmid vector (pmk16) containing SV40 DNA using a liposome-mediated method. The viral DNA was from a replication-defective, origin-minus, temperature-sensitive mutant strain (A58). One clone, designated PE-1, has been propagated for over 120 passages. PE-1 cells grown at 33C (33C cells) exhibit spindle-shaped morphology; when cultured at 40C (40C cells), they took on a polygonal or spherical shape. Morphology of 40C cells returned to the spindle shape after culture flasks were shifted back to 33C. During a 2-week period, 33C cells propagated approximately 30-fold faster than 40C cells, whereas protein concentration was higher in 40C cells. Southern blot analysis of PE-1 cells demonstrated successful integration of the ts-SV40 DNA sequence into the porcine endometrial cells, possibly at multiple sites. The presence of cytokeratin on PE-1 cell membranes was shown by immunocytochemical studies, suggesting that the PE-1 cell clone was of epithelial origin. Reverse phase (RP)-HPLC analysis of PE-1 cell extract indicated that the majority of immunoreactive beta-endorphin (ir-BEND) eluted with a hydrophobicity similar to that of synthetic BEND and alpha-N-acetylated BEND (Nac-BEND). These results demonstrate that a porcine endometrial cell line has been established, and that this cell line possesses characteristics of temperature sensitivity in cell morphology, growth rate, and protein synthesis.
Animals↗
Effect of propranolol on calcium, phosphorus, and magnesium metabolic disorders in Graves' disease.
The purpose of this study was to determine whether propranolol alone can improve mineral metabolic disorders in thyrotoxicosis. Ten Graves' disease patients and 11 normal age- and sex-matched controls participated in the study. In the untreated Graves' patients, serum levels of calcium (Ca), calcium x phosphorus product (Ca x P), urinary Ca, phosphorus (P), magnesium (Mg), and hydroxyproline (Hp) were higher than in control subjects (P less than .05), intestinal Ca absorption was lower than in control subjects (P less than .05), and Ca, P, and Mg balance were negative (P less than .05). After 40 mg propranolol four times per day (qid) for 28 days, serum triiodothyronine (T3) had decreased (P less than .05), serum reverse triiodothyronine (rT3) increased (P less than .05), serum thyroxine (T4) remained unchanged (P greater than .05), serum Ca and urine Ca and Mg decreased (P less than .05), intestinal Ca absorption increased, Ca balance was corrected, and P and Mg balance was improved (P less than .05). Our results indicate that propranolol can improve the metabolic disorders in addition to the symptomatic manifestations of Graves' disease. The mechanism responsible for the improved mineral balance is unclear, but may be related to beta-adrenergic blockade, increased membrane stability, or a decrease in the thyrotoxic state caused by the therapeutically induced decrease in serum T3.
Adolescent↗
Endometrial immunoreactive beta-endorphin increases during mid-estrous cycle and early pregnancy in gilts.
Immunoreactive (ir) beta-endorphin (BEND) was recently identified in porcine uterine fluids. In the study reported here, we examined the hypothesis that porcine endometrium serves as a source of uterine fluid ir-BEND during the estrous cycle and early pregnancy. Endometrial ir-BEND was chromatographically characterized, sites of ir-BEND synthesis were immunocytochemically localized, and concentrations of endometrial ir-BEND during the estrous cycle and early pregnancy were measured. Sephadex G-50 chromatographic profiles of endometrial extracts from Day 15 of the estrous cycle revealed three distinct peaks of ir-BEND, with the first peak occurring near void volume and the second and third peaks coinciding with standard porcine beta-lipotropin and standard porcine BEND, respectively. Reverse-phase HPLC C18 chromatographic profiles indicated that endometrial ir-BEND contained both standard BEND and alpha-N-acetylated BEND. Immunocytochemical studies demonstrated ir-BEND in the surface and glandular epithelial cells of the endometrium, with immunostaining most prominent in the apical portion of epithelial cells. Concentrations of ir-BEND in endometrial tissues were higher on Days 14-15 than on Days 8-12 during the estrous cycle and pregnancy (p less than 0.05); however, values were not different in pregnant and cyclic gilts. Biochemical and immunocytochemical evidence supports our hypothesis that ir-BEND present in uterine fluids is derived from the endometrium. The increase in endometrial ir-BEND concentration during Days 14-15 in cyclic and pregnant gilts indicates that ovarian steroids may influence the synthesis of endometrial ir-BEND.
Animals↗
An X-linked human collagen transgene escapes X inactivation in a subset of cells.
Transgenic mice carrying one complete copy of the human alpha 1(I) collagen gene on the X chromosome (HucII mice) were used to study the effect of X inactivation on transgene expression. By chromosomal in situ hybridization, the transgene was mapped to the D/E region close to the Xce locus, which is the controlling element. Quantitative RNA analyses indicated that transgene expression in homozygous and heterozygous females was about 125% and 62%, respectively, of the level found in hemizygous males. Also, females with Searle's translocation carrying the transgene on the inactive X chromosome (Xi) expressed about 18% transgene RNA when compared to hemizygous males. These results were consistent with the transgene being subject to but partially escaping from X inactivation. Two lines of evidence indicated that the transgene escaped X inactivation or was reactivated in a small subset of cells rather than being expressed at a lower level from the Xi in all cells, (i) None of nine single cell clones carrying the transgene on the Xi transcribed transgene RNA. In these clones the transgene was highly methylated in contrast to clones carrying the transgene on the Xa. (ii) In situ hybridization to RNA of cultured cells revealed that about 3% of uncloned cells with the transgene on the Xi expressed transgene RNA at a level comparable to that on the Xa. Our results indicate that the autosomal human collagen gene integrated on the mouse X chromosome is susceptible to X inactivation. Inactivation is, however, not complete as a subset of cells carrying the transgene on Xi expresses the transgene at a level comparable to that when carried on Xa.
Animals↗
New bone formation in the in vivo implantation of bioceramics. A quantitative analysis.
Two kinds of synthetic biomaterial, porous tricalcium phosphate (PTCP) and magnetic porous tricalcium phosphate (MPTCP) ceramic granules were implanted in rat femur. In the period of 4 months, the assessment of serial histological sections, scanning electron microphotographs and quantitative analysis of bone formation in the sections showed that both ceramics are biocompatible and degradable in vivo. More new bone formation occurred in the MPTCP group. Endochondral ossification was seen in both groups. The quantitative analysis in this study is reliable, and may be suitable to the similar experimental models.
Animals↗
[Preparation and utilization of BrdU antiserum. II. Detections of BrdU labelling chromosome and DNA using BrdU antibody technique].
Utilizing highly specific BrdU antiserum, via the second antibody conjugated with enzyme and DAB-H2O2 or AEC-H2O2 colorific methods, the BrdU marked regions of CHO and human chromosomes were detected clearly. The BrdU-DNA prints (pg) on nitrocellulose membranes were also showed with similar methods.
Animals↗
[Effects of several primary factors related to establishment of mouse embryo pluripotent stem cell lines (ES cell line)].
Several primary factors related to establishment of mouse embryo pluripotent stem cell lines (ES cell lines) have been tested comparatively. The results of 825 embryos indicated: (1) Delayed blastocysts were more conducive than normal ones to proliferation of inner cell mass (ICM). (2) Culture medium DMEM with 4 500 mg/l glucose was more advantageous than DMEM with 1000 mg/l glucose to the attachment of explanted blastocysts and to the proliferation of ICM. (3) DMEM without sodium pyruvate was beneficial to appearance of ES cell colonies and to their growth. (4) Feeder layer prepared from the primary culture of mouse embryos had very good effects to the growth and culture of ES cells.
Animals↗
[Evaluation of preventive measures for osteoporosis].
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Aged↗
Potentiation of cyclic adenosine monophosphate production by thrombin in the human erythroleukemia cell line, HEL.
The human erythroleukemia cell line (HEL) has been used as a model system for studying signal transduction processes as they might relate to platelet/megakaryocyte function. We were interested in examining the role of thrombin in the regulation of adenylyl cyclase in this cell line. As opposed to its predominantly inhibitory effects on cyclic AMP production in platelets or in membranes from HEL cells, our initial experiments in intact HEL cells revealed that thrombin markedly potentiated the cyclic AMP response to prostaglandin E1 (2.9 +/- 0.2-fold), prostacyclin (1.9 +/- 0.2-fold) and carbacyclin (2.5 +/- 0.5-fold), measured either by radioimmunoassay or by the [3H]adenine preloading procedure. Thrombin, although ineffective alone, also potentiated cyclic AMP production stimulated by vasoactive intestinal peptide (1.6 +/- 0.2-fold), cholera toxin (3.0 +/- 0.6-fold) and AIF4- (2.3 +/- 0.6-fold), but not by forskolin (0.9 +/- 0.1-fold). The thrombin effect 1) produced an increase in the efficacy of the prostaglandins with no change in potency; 2) was long-lived; 3) required the proteolytic activity of thrombin; 4) was insensitive to pertussis toxin; and 5) was at least partially mimicked by trypsin, extracellular ATP and UTP, platelet activating factor and activators of protein kinase C. Down-regulation of protein kinase C or pre-exposure to the protein kinase inhibitor staurosporine blocked the potentiating effect. Together, these results suggest that in HEL cells, the mechanism of thrombin potentiation of cyclic AMP production may involve alterations in the interaction between stimulatory guanine nucleotide binding protein and the catalytic subunit of adenylyl cyclase, possibly involving protein kinase C-mediated phosphorylation.
Adenylyl Cyclases↗
Rabies in China: recommendations for control.
Reviewed are the results of 15 years' experience with rabies at You-An Infectious Disease Hospital, Beijing, China. The purpose of the study was to determine whether there are any epidemiological or clinical features of rabies that are unique to China and which might be important in developing a strategy to control it. During the period under study, 64 patients with rabies were admitted to You-An Hospital. Exposure to dogs was associated with 61 cases, two involving the handling of dog carcasses that were being prepared for meals. All of the exposures occurred in rural areas, and none of the patients received adequate prophylaxis. Patients with proximal sites of exposure and with severe injuries developed rabies after short incubation periods (P less than 0.05, and P less than 0.02, respectively). Failed vaccination was also associated with a short incubation period (P less than 0.05). Haematemesis occurred in 20 patients and was associated with shorter incubation periods (P less than 0.02), facial exposure sites (P = 0.021), and severe injuries (P = 0.047). A strategy to control rabies in China should include efforts to educate the public about handling the carcasses of stray dogs, in addition to the currently recommended strategy of controlling the dog population and of vaccinating domesticated animals.
Adolescent↗
[Growth regularities of yujin (Curcuma longa L.)].
Based on the characteristics of leaf, root and tuber growth as well as dry matter accumulation, the ontogeny of Yujin is divided into 4 stages. The growth and correlation among the leaf, root and tuber as well as accumulation of essential oils and curcumins have also been studied.
Curcumin↗
[Effect of baoyuantang on erythroid progenitor and clinical observation in chronic renal failure patients].
The erythroid colony formating ability and the therapeutic efficacy were investigated in patients with chronic renal failure (CRF) before and after treatment with baoyuantang. The results showed that the number of CFU-E, BFU-E of bone marrow was found lower in CRF patients than that in normal controls, and there was inhibitory action in the CRF serum on the number of CFU-E, BFU-E of normal human bone marrow. After treatment with baoyuantang, the amount of Hb was increased and creatinine decreased in CRF patients, and its effective rate reached 83.3%. The above-mentioned inhibitory action in CRF serum was also reduced post-treatment. The results indicated that baoyuantang could increase the ability for proliferation and differentiation of erythroid progenitor and adjust anemia in CRF patient.
Adult↗